Assisted Repor: Artificial Insemination (A.I.) and Embryo Transfer (E.T.)

Question 1

Artificial insemination

Answer 1

Bovine, Equine, Canine, Ovine, and other species

Sperm is collected from the maile and placed into the female reproductive tract to join with a naturally ovulated egg

Question 2

Sex-Selected semen

Answer 2

Bovine and Equine

Sperm is sorted by flow cytometry into predominantly X-bearing sperm and predominantly Y-bearing sperm aliquots

the sorted sperm is then used for AI

Question 3

Embryo Transfer

Answer 3

Bovine and equine most commonly

AI or natural service is used to inseminate a female

The resulting embryo(s) are collected prior to the time of “hatching” from the zona pellucida and transferred to the different female (recipient) ofr the rest of gestation

Question 4

In vitro Fertilization

Answer 4

Becommong Common

A single sperm cell is inserted into an egg in a laboratory environment

Embryo is cultured on artificial bedia of several days and if the embryo proceeds to the blastocyst stage - it is transferred inot a recepient (donor or another female) for the rest of gestation

Question 5

Gamete Intra-Fallopian Transfer (GIFT)

Answer 5

Mostly experimental

An egg is aspirted from a donor female’s ovary prior to ovulation and then planced in the uterine tube of a recipient

THe recipent is bred naturally or via AI

Question 6

Embryo-splitting

Answer 6

Bovine and Equine

Identical twins (or more) can be produced by splitting a day -6 embryo (blastocyst before hatching from ZP) – and placing the fragments into empty ZP shells and then transferring them into recipients

Question 7

Cloning (Nuclear transfer)

Answer 7

Bovine, ovine, canine, and equine

Multiple genetically identical animals can be produced by taking cells from an embryonic blastocyst and implanting them into enucleated oocytes which are transferred to a recipient

Question 8

Cloning (Somatic Cell Nuclear Transfer)

Answer 8

Ovine, canine, bovine, and equine

Multiple genetically identical animals can be produced by taking an adult cell and “re-programming” it to allow differentiation into an entire organism when implanted into enucleated eggs

Question 9

Artificial Insemination

Advantages

Answer 9

Genetic Improvement - Widespread use of males likely to produce valuable offspring (based on the sire’s performance) or proven to have valuable offspring (based on many documented offspring)

Can use males form anywhere in the world

Don’t have to have as many males

Remove/reduce risk of venereal disease

Question 10

Artificial Insemination

Disadvantages

Answer 10

Skill of estrus detection

Trained insemination - skill that must be hired or learned

Time and skill to commit to herd/animal reproduction

Increased cost per pregnancy

Question 11

Collecting Semen for AI

Answer 11

Semen can be purchased form commercial suppliers and directly from males owners

Semen can be collected on-site and used to mate many more females and at decreased risk of injury than via natural service

Bull, Dog, and stallion semen is frequently frozen - other species primarily use fresh or chilled semen

THe semen of some males does not freeze well - this is not necessarily related to fertility in natural service

Question 12

Collecting semen for AI

Artificial Vagina

Answer 12

Males that are trained to mount a dummy or teaser animal

Bulls, Stallions

Stallion (temperature / pressure ejaculator) - AV has a jacket that is filled with warm water

Bulls (Pressure ejaculator) - AV does not need a warm water jacket

Question 13

Collecting semen for AI

Electroejaculation

Answer 13

Males that are not trained to mount a dummy or teaser animal

Bulls, bucks, Wildlife

Specialized probe placed into the rectum applies a series of low-voltage pulses to the pelvic nerves involved in erection and ejaculation

Question 14

Collecting Semen for AI

manual Stimulation

Answer 14

Pressure ejaculators - massage the penis

Dogs, boars, whales

Collection tube rather then true AV

Question 15

Collecting Semen for AI

DO NOT USE

Answer 15

latex, plasitc, and lubricants can be detrimental to sperm viablility

Limit their use and contact time

Question 16

Cryopreservation of sperm

Adequate fertility post-thaw requires:

Appropriate diluents / extenders

Answer 16

Non-penetrating cryoprotectant

Penetrating cryoprotectant to protect from ice cyrstal formation

Buffer to prevent pH change

Sugar to provide an energy source

Salt to provide proper osmolality

Antibiotic to kill pathogens

Question 17

Cryopreservation of sperm

Adequate fertility post-thaw requires:

Appropriate Sperm Dilution

Answer 17

Bulls: 20 million sperm per dose - assume 50% live sperm post-thaw

Can use a lot less - but must thaw and place semen very well

Question 18

Cryopreservation of sperm

Adequate fertility post-thaw requires:

Appropriate Cooling and Thawing Speed

Answer 18

Rapid Cooling - but in steps

Add some extender ingredients and cool from 102F to 40F - refrigeratation temp

Add cyroprotectants and package into 0.5 mL straws - hold at 40F for a few hours

Freeze in liquid nitrogen vapor at -238F

Store in liquid nitrogen until used -320F

Question 19

Cryopreservation of sperm

Adequate fertility post-thaw requires:

Rapid Warming

Answer 19

Semen thawed at 95F for 12 seconds to one minute had best post-thaw viability

Semen thawed at 68F for 12 seconds to one minute has acceptable post-thaw viability

Semen thawed at 41F water bath or in a shirt pocket has lowest post-thaw viability

Question 20

Artificial Insemination

Detection of Estrus

Answer 20

Used to estimate timing of ovulation

Difficult - great species-to-species viariability and animal-to-animal variability within a species

Question 21

Artificial Insemination

Timing of Insemination

Canine

Fresh Semen

Answer 21

Ovulation occurs 1 to 2 days after LH surge

Wait longer to breed after ovulation

Fresh semen: lives 5-6 days in uterus

Breed 4-6 days after LH surge

Question 22

Artificial Insemination

Timing of Insemination

Canine

Chilled Semen

Answer 22

LIves about 1-3 days after ejaculaiton

Will lose at least 1 day in shipping

Breed 4-5 days after LH surge

If breeding 2 times, breed at days 3 and 5; 4 and 6 post LH peak

Must plan ahead - order semen when she ovulates

Question 23

Artificial Insemination

Timing of Insemination

Canine

Frozen semen

Answer 23

Lives about 12 hours in uterus

Breed 5-6 days after LH surge

no Problem with shipping at the last minute - have the semen on hand far in advance of breeding

Intrauterine insemination - surgical or transcervical catheterization

Question 24

Artificial Insemination

Timing of Insemination

Bovine

Answer 24

Almost always utilize frozen semen

Successful breeding with frozen semen since 1940s

Fertility using forzen semen in equal to natural service

Semen is placed in the uterus - pipette is passed through the cervix

Breed approximately 12 h after first detection in esturs

Question 25

Artificial Insemination Timing of Insemination Bovine Frozen straws

Answer 25

Handle with care Canister in the liquid tank contains several “canes” filled with several “straws” of semen The straw you are removing is not the problem - be careful not to let the staws around it to warm up Once thawed - do not let the straw cool

Question 26

Artificial Insemination Timing of Insemination Equine

Answer 26

Fresh semen us used immediatley does not have to be extended Fresh semen used 10 minutes or longer after collection should be extended Cold shold causes plasma membrane damage and acrosome changes - reduced fertility Extended semen used within 6-12 horus can be stored at room temp Removing seminal plasma delays capacitation - extends life of sperm for transport To maintain viability for longer than 12 hours - cool to 40-45F over a 2-3 hour period Ship in container designed to maintian steady temperature

Question 27

Artificial Insemination Timing of Insemination Equine Chilled semen

Answer 27

Do not expect to remain viable longer than 60-72 hours Inseminate mares within 24 hours prior to ovulation 1.8mg deslorelin acetate cna be used in mares with a \>35mm follicle to induce ovulation ~85% will ovulate w/n 24-48 hours Sucromate equine is in FDA appreved frug contianing deslorelin and therefore it is not legal to use a compounded deslorelin

Question 28

Artificial Insemination Timing of Insemination Ovine

Answer 28

Ram effect is utilized to aid in synchronization AI si not commonly done in ewe becuase it is dufficult to detect estrus AI is not commonly done in the ewe becuase it is difficult to pass a catheter through the cervix

Question 29

Artificial Insemination TIming of Insemination Ovine

Answer 29

Cervical AI - speculum used to visualize the cervical os and fresh or chilled semen is deposited in the cervix

Question 30

Artificial Insemination TIming of Insemination Ovine

Answer 30

Transcervical AI - special forceps are used to pull the cervix into the vagina where another specialized scope is used to guide a catheter through the cervix

Question 31

Artificial Insemination TIming of Insemination Ovine Laparoscopic AI

Answer 31

surgical endoscopic approach to deposit semen directly into uterus