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Protocol for breeding via live cover:  Before a mare is allowed to enter a shed for breeding, a veterinary certificate should be required by a stallion station to confirm that a mare has no physical signs of genital tract infection and that a recent uterine culture yielded no significant growth of potentially pathogenic bacteria.  Definitive identification of the mare should be required.  All regulatory requirements must be followed.  To verify requirements in your area, please check with your state/provincial animal health office (see State Veterinary Offices in the AAEP Resource Guide and Membership Directory).  A current negative test result for Equine Infectious Anemia virus (i.e. either a Coggins test or EIA ELISA) and a Certificate of Veterinary Inspection (CVI) may also be required. 

Mares may also be required to have received specific vaccine(s) during an approved interval prior to visiting the breeding shed.  Depending on the initial breeding status of the mare (maiden, foaling, barren), the stallion station may require an additional culture for each heat period in which the mare is bred (i.e., cultures carried out more than 30 days previously may not be acceptable).  Negative cultures and complement fixation (CF) tests for Taylorella equigenitalis/asinigenitalis may be required prior to breeding of imported mares. 

A vaginal speculum examination, using a sterile (preferably disposable) equipment, is often an integral part of the pre-breeding evaluation in a live cover program, but may not be required as part of the veterinary health certificate.  Breeding shed requirements may vary between maiden, foaling, barren and imported mares. 

Disposable gloves should be worn by the person cleaning the stallion’s penis and directing it into the mare’s vagina.  These gloves should be discarded into a covered trash receptacle prior to contacting other surfaces such as areas in the breeding shed or equipment used to handle the mare or other stallions.  Disposable gloves should also be used by personnel holding the tail and/or breeding roll, and lead rope (and twitch if used). If a breeding roll is used during a live cover, the roll should be covered with a disposable plastic sleeve which is changed between horses.  If a twitch is used to restrain a mare during a live cover, the twitch should be disinfected prior to use on another horse.  Before each breeding session, all facilities used to handle and breed mares, as well as the breeding shed itself, should be cleaned and disinfected as necessary.

Protocol for maintenance of artificial vaginas:  Appropriate cleaning and storage of artificial vaginas will aid in controlling horizontal transmission of venereally transmitted diseases.  Personnel cleaning an artificial vagina should wear disposable gloves/sleeves and change these between each cleaning procedure.  One accepted method of cleaning an artificial vagina, is provided below: 

  1. Rinse the artificial vagina liner (inside and outside) thoroughly in hot (greater than 50-degrees Celsius/122-degrees Fahrenheit) running tap water and use a clean large bottle-, flask- or beaker-brush (brush should be cleaned with hot water and 70% isopropyl alcohol between uses) to aid in removal of any particulate matter that is adhered to the latex rubber.  Since soaps or disinfectants (other than alcohol) may permeate the latex rubber and subsequently leach into semen, their use is not recommended.
  2. Rinse the artificial vagina with deionized or distilled water to remove any impurities in the tap water that could negatively impact sperm function.
  3. Rinse the artificial vagina with 70% isopropyl or ethyl alcohol, ensuring the entire latex liner is covered.  Rinsing can easily be accomplished with a squirt or spray bottle that is designated for this purpose.
  4. Hang the artificial vagina in a clean covered cabinet for drying and storage (protected from UV rays or natural sunlight to avoid drying and cracking of the liner over time).
  5. Thoroughly rinse the sink used for cleaning artificial vaginas with tap water after each use, then wipe or rinse sink with 70% alcohol. 

It would be ideal to have a latex artificial vagina liner dedicated for each individual stallion.  It would also be ideal to clean used artificial vagina liners in a sink separate from the one used during filling of liners.  Alternatively, disposable artificial vagina liners could be used for each stallion.  Plastic disposable liners can be used to collect semen from stallions to reduce transmission of venereally transmitted diseases if the stallion will tolerate its use. 

Latex artificial vaginas that show material breakdown (cracks, thin spots, sticky spots or black areas) should be discarded and replaced.   Between uses, the case/cover of the artificial vagina should be thoroughly cleaned with hot water, then sprayed or rinsed with 70% isopropyl alcohol and allowed to air dry. 

Protocol for maintenance of breeding phantoms:  Contaminated breeding phantoms have a high potential for horizontal transmission of venereal pathogens.  It is advisable to thoroughly cover the back end of the breeding phantom (the portion that comes in contact with the stallion’s genitalia) with a disposable plastic wrap prior to use.  The wrapping material should be fitted properly to the breeding phantom such that it will not easily come off during the semen-collection process.  The wrapping material should be removed after each use and discarded.  The cover of the breeding phantom should then be cleaned after each use.  Personnel should wear disposable gloves/sleeves when cleaning a breeding phantom and the gloves should be discarded after use.  The phantom can be washed with soap and water, if visible debris is present, and then disinfected by applying 70% alcohol.  To avoid damage to semen during collection, disinfectants should only be used on the breeding phantom if any residue will not contaminate the semen during collection and impair its quality (once dried, alcohol leaves no residue, in contrast to some other disinfectants).  Reusable and washable covers can be made for individual stallion use.  Where only farm horses previously confirmed free from venereal disease are involved, modification of this cleaning regimen can be considered.

Protocol for AI: Routine evaluation of the semen prior to insemination may reveal signs that an infectious agent may be present (e.g., numerous neutrophils, contaminating debris).  Further diagnostic testing should be done if deemed necessary. 

It is recommended that all semen be diluted in a semen extender containing appropriate antimicrobial drugs (AMD), which may limit bacterial growth and transmission of potentially pathogenic bacteria by AI.  Most commercial equine semen extenders contain AMD, or an AMD can be added to the base ingredients.  In some cases, AMDs are not present in commercial extenders at desirable concentrations, in which case further AMD may be added. Be aware that, bacterial or fungal pathogens may survive in extended semen containing AMD and might still result in infection of a mare following insemination. 

Consider the contents of a transported-semen shipping container as potentially contaminated.  While there is no way to recognize the presence of CEMO or EAV in transported or frozen semen (other than culture or PCR testing), be aware that both diseases have been transmitted by breeding with infective semen from shedding stallions, and even recipient mares have been infected with EAV following transfer of embryos derived from donor mares bred with infective semen. Wear disposable gloves when opening a shipping container and handling packages of shipped semen.  Clean all surfaces of the laboratory area in contact with the semen container and semen packages. Wash hands thoroughly with soap and water after removing disposable gloves.   

Use sterile disposable equipment for insemination, including syringes, insemination pipette, and use sterile tubes or individual sachets of obstetrical lubricant and obstetrical sleeves.  Discard all disposable equipment promptly and properly.  Wash hands with soap and water once all procedures are completed. If possible, clean and disinfect entire area, including examination equipment and flooring, after breeding/insemination procedure(s) is/are completed.