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Health Testing

Image00169Health Testing
“Clearances” – Varies from breed to breed. Some of the more common tests include:

  • CERF

  • Hip Dysplasia Evaluation - OFA/PennHIP

  • Elbow Dysplasia - OFA

  • Luxating Patellas - OFA

  • Cardiac Evaluation - OFA

  • Thyroid Function - OFA

  • Serologic testing

  • Brucella canis RCAT – high sensitivity, low specificity

  • False positives common

  • Confirm with AGID test

  • Herpesvirus testing


Prebreeding examination
  • General physical examination

  • Reproductive examination – digital vaginal examination, examination of external gentalia

  • Verification of protection from infectious viral disease (immunizations are current or vaccination titers are protective)

  • Herpesvirus testing

  • Testing to ensure candidates are free from parasitic infections (heartworm disease and intestinal parasites

  • Prebreeding vaginal cultures


What – Type of breeding


Natural mating
  • Dog and bitch are both fertile

  • Dog and bitch are not separated geographically

  • Both are available at the appropriate time

  • Neither have a physical contraindication (arthritis, neurologic disease, vestibulovaginal band or stricture, size incompatibility)

  • Both exhibit receptive behavior


Image00173Artificial insemination with fresh semen
  • Dog and bitch cannot or will not copulate normally

  • Bracheocephalic breeds

  • Social incompatibility – dominant female will not stand to be mounted

  • Bitch has a congenital vaginal anomaly

  • Dog is unable to intromit

  • Size incompatibility – seen more in designer breeds

  • Documentation of semen quality is desired


Artificial insemination with fresh chilled extended semen
  • Circumvents the requirement for the dog’s presence at the breeding

  • Circumvents the need for the semen collection to be performed on the same day as the breeding

  • Permits breeding of individuals located distantly from each other


Artificial insemination with frozen semen
  • Allows indefinite storage of a dog’s genetics

  • Permits siring of litters with sperm cells from deceased dogs

  • Permits transport around the world

  • Post-thaw longevity of frozen semen is very short (approx. 12 hours)

  • Requires accurate ovulation timing

  • Requires intra-uterine sperm cell deposition


Plan the breeding when your client is the bitch owner:


1. Plan ahead!
2. Make sure you have all the supplies you will need (insemination pipettes, nonspermacidal lubricant, semen extender, etc.)
3. Confirm with your commercial laboratory that progesterone assays are of the desired type, are run daily, and are results are reported back within 24 hours.
4. Don’t commit to organizing and performing an artificial insemination (AI) if you will not be available. This includes weekends and holidays.
5. Record the contact information for all parties involved (bitch owner, dog owner, dog owner’s veterinarian, handler, guardian, etc.)
6. Avoid a “Garbage in, garbage out” breeding – confirm that the male is currently fertile and in good health.
7. Plan the breeding with young, healthy fertile individuals. Since this is almost always out of the clinician’s control, special measures may need to be taken if either the bitch or dog is aged or known to be subfertile. For example, fresh chilled semen of marginal quality may necessitate surgical rather than vaginal insemination to achieve pregnancy.
8. Make sure that the bitch to be bred is healthy and able to withstand the rigors of pregnancy and puppy rearing. Check heartworm status, thyroid function, etc.
9. Review the bitch’s nutrition with the owner. It is best to have the bitch in a “weight-gain” status at the time of conception. Have the owner supplement the bitch’s diet with B vitamins and omega fatty acids.
10. Instruct your client to contact the dog owner and confirm that the dog will be “ready, willing and able” to provide the necessary services when the time comes.
11. Be prepared to “change gears.” An anticipated vaginal insemination may end up being a surgical insemination if the semen quality is marginal upon its arrival.
12. Recommend that the bitch owner always have a Plan B ready to be implemented in the event that the scheduled breeding encounters a “hitch”. (Example, Dog A is away at a show and cannot be
collected at the proper time or semen arrives “DOA.”)
13. Recommend that your client request confirmation of current negative Brucellosis status on the dog. I (arbitrarily) recommend that the dog have a negative test result for Brucella canis within:
  • 30 days, or

  • since his last natural breeding, or

  • since his last congregation with other dogs, whichever is the most recent. Be aware that Brucellosis, often thought to be only a sexually transmitted disease, can also be transmitted by casual contact.


Perform Ovulation Timing – stay tuned to our next hour on “It’s all in the timing!”


Schedule the Inseminations
1. Typically with vaginal AIs, two inseminations are performed 48 hours apart during the bitch’s period of peak fertility.
2. Inseminations are performed 2 and 4 days post-ovulation.
3. Confirm that the dog owner and the dog owner’s veterinarian have been contacted and that semen collections are scheduled for the day preceeding the day of insemination in each of the
two inseminations.
4. On the days of scheduled semen collection, call the collecting veterinarian’s office and obtain the tracking numbers for the shipments.

Evaluate the Semen
1. When fresh chilled semen package arrives, it should be opened immediately. Attention should be paid to the "impression of coldness." The ice packs should be at least cold, if not still frozen.
2. The tube containing the semen should be removed from the packaging material (usually newsprint). The tube should contain the extended semen in a liquid state. Unfortunately, occasional mishandling by the shipping company or by the shipper placing the semen package in a non-pressurized compartment of the airplane will cause the sample to arrive frozen. The freezing kills the sperm cells and renders the sample useless.
3. Read the accompanying semen shipping form and note any special handling instructions that may be included.
4. One drop of the sample should be placed on a warmed microscope slide. The rest of the sample should be refrigerated. Allowing the chilled sample to warm to room temperature only allows the
sperm cells to speed up, using precious energy and shortening their life span.
5. Semen arriving looking “DOA” may still be viable. Some extenders (eg., Clone) render the sample immotile and additional of an activator buffer is required to restore sperm cell motility.
Warming or transferring apparently lifeless semen into fresh extender may bring it back to life. Perform test manipulations on only a single drop of semen. As the semen drop warms, side to
side motility becomes noted. The continued warming eventually shows the cells to have achieved a normal forward progression. As the drop warms on the slide, read the accompanying paperwork
with the semen collector's evaluation of the semen quality. Time of collection and post-collection motility should be studied. The semen drop is then analyzed and compared to the collector's
evaluation.
6. If no motility is noted after 15 minutes, the sample is most likely non-viable. If this occurs, the collector of the semen should be contacted to determine, if possible, the cause of the semen's demise. In other cases where only partial semen recovery is noted, the inseminator must use judgment based on the concentration of the semen, estimated total spermatozoa numbers and
the percent recovered. It may also be necessary to alter the insemination method to that of an intra-uterine deposition of the fresh chilled sperm to further reduce sperm cell stress and to aid its arrival at the fallopian tubes.
7. The refrigerated fresh chilled sample need not be warmed to room temperature or body temperature before insemination. Having the sample in the uterus as it warms makes maximum
use of the conserved energy. All fresh chilled semen samples are handled in a similar manner, however, many different commercial companies sell packaging kits and extenders. One should
always read their instructions for any specific handling recommendations before using the semen.

Perform the Insemination
1. Most often, fresh chilled semen is inseminated vaginally. The pregnancy rate for vaginal inseminations using good quality semen should be at least 80%. Surgical insemination or TCI
(transcervical insemination) can be performed to increase pregnancy rate. Deposition of the semen directly into the uterus improves pregnancy rate when chilled semen quality is marginal.
Uterine deposition of semen also improves pregnancy rate in very small and giant breeds of dogs, which are reported to have less success with vaginal inseminations.
2. To perform a vaginal insemination, the pipette is introduced through the dorsal vulvar comissure, directed dorsocranially through the vagina until it enters the pelvic vagina. Then the pipette is directed cranially as far as possible, ideally to the external cervical os. The bitch’s hind end is elevated to a 45 degree angle while the semen is injected through the pipette. The dorsal vaginal wall is gently “feathered” for a moment or two while the hind end remains elevated for 10 minutes. Care is taken not to place pressure on the caudal abdomen during this time.
3. Recent reports indicate that elevating the hind end for longer than one minute offers no advantage over a one minute elevation.
4. Save the semen tube for DNA verification of paternity should that become necessary.

Planning the breeding when your client is the dog owner:


1. Plan ahead! Set up accounts with Fed Ex and UPS. Become a “known shipper” with several airlines so that you can offer “counter to counter” shipments for your clients. This involves a fee
and site inspection. If possible, set up “Same Day” service with Fed Ex. This also involves a site inspection. Set up an account with a local courier service so that you can provide delivery of the collection from your facility to the airport, if need be.
2. When working with breeder clients, instruct them to contact you one month before the anticipated breeding.
3. Make sure you have all the supplies you will need: collection cones, nonspermacidal lube, semen extender, 15 ml conical tubes with crew caps, parafilm, Spermacue™ or hemacytometer,
shipping boxes, semen evaluation forms, shipping forms, etc.
4. If the dog has never been collected and shipped before, a trial collection and “chill check” is advisable at least a week before to the anticipated shipping date. This ensures that the dog can be collected and that his semen quality is maintained after extending and chilling. Rarely, a dog can be “extender specific.”
5. If the dog has not been collected recently, a trial collection verifies that semen quality has not deteriorated. Many dog owners also request a trial collection to “flush the rusty pipes.”
6. Obtain contact information from all parties involved. Also find out how (vaginal, TCI, or surgical) the insemination will be performed. Insemination method determines some aspects of the semen processing.

Schedule the Collections
1. Schedule collections to minimize the collection to insemination time interval. Typically, the dog is collected for overnight shipment and the insemination is performed the next day.
2. Shipment usually involves an overnight FedEx service. Call Fed Ex and schedule the pick up as soon as you know the collection date.
3. Verify that you have the correct name, address and phone number for the recipient. Usually the collection is shipped directly to the inseminating veterinarian. Rarely, the collection is shipped to the bitch owner’s residence or workplace.

Perform the Collections
1. Remove the semen extender from the freezer well ahead of time. The extender and the ejaculate must be at the same temperature when they are combined.
2. Use an estrous teaser bitch whenever possible. Spermatozoa number and quality are maximized when a teaser bitch is available. As much as a four-fold increase in spermatozoa number can be
realized when using a teaser bitch. General guidelines for adequate semen quality include >70% motility, >70% morphologically normal spermatozoa, few inflammatory cells, and a normal sperm
count. Sperm count should approximate 10 million spermatozoa per pound of body weight. A collection of at least 250 million “good ones” is desirable.
3. If collected sperm numbers are marginal (<250 million sperm), wait 45 minutes and collect the dog a second time. If the total number of sperm cells remains low, recommend that the dog
owner allow you to call the bitch owner to confirm that he or she still wants to have the suboptimal collection shipped.
4. When collecting the dog, collect only the sperm-rich second fraction of the ejaculate. Excess prostatic fluid is deleterious to semen quality. If excess prostatic fluid is present, centrifuge the ejaculate, decant off the supernatant and resuspend the sperm pellet in an appropriate extender. Follow semen processing directions carefully. Many extenders are available, each with its own handling instructions.

Process and Package the Semen
1. When extending semen, it is important to know how the insemination will be performed. Surgical insemination requires much less total volume than does vaginal insemination. For routine vaginal insemination, the total volume should be appropriate for the size of the bitch. Generally, 2 to 3 mls is adequate for small breeds, 4 mls for medium breeds, 6 mls for large breeds and up to 10 mls for giant breeds.
2. The sperm rich fraction or sperm pellet is resuspended to obtain an appropriate total volume. The sperm to extender volume ratio must be at least 1:1. A ratio of 1 part ejaculate to 3 parts
extender is common.
3. Send the extended semen sample in a conical shaped centrifuge tube with a screw cap.
4. LABEL THE TUBE. Identify the shipment with the dog’s name (call and/or registered name) and breed.
5. Make sure the tube does not leak. It is also a good idea to wrap the cap with a layer of parafilm. Placing the tube inside a whirl pack or sealable plastic bag allows for recovery of semen should a leak or breakage occur.
6. A number of shipping containers are available commercially. Shipping kits can be obtained from Camelot Farms, Clone, ICSB, Minitube, Reproduction Resources or Synbiotics. Equitainers can
also be used. Styrofoam boxes work adequately in most instances. In Europe, insulated Thermos-type containers are popular.
7. The bagged tube is placed in the box along with 1 or 2 frozen gel pack or Kool-It brick. The tube should not be placed in direct contact with the frozen coolant. Make sure that the tube is
separated from the brick by at least 4 to 6 layers of paper towel or newspaper. The tube can be wrapped with an insulating material or the tube can be placed in a smaller Styrofoam box. More
newspaper (sufficient to fill the dead space) is packed between the brick and the tube. Alternatively, one refrigerated brick can be placed between the frozen brick and the tube, still
using newspaper to fill dead space.
8. SEND QUALITY ASSESSMENT DATA. Put a copy of the semen evaluation and processing form, and the inseminating instruction sheet in a baggie and place it in the shipping box.
9. Seal the shipping container and attach the completed Fed Ex or UPS form. When dealing with weekend inseminations, make sure that package can be delivered on Saturday mornings. There
are often special boxes on the shipping form that need to be checked for AM and Saturday delivery.
10. Call and give the FedEx or UPS tracking number of the shipment to the inseminating veterinarian.
11. The cost of semen collection and shipping is typically the bitch owner’s responsibility. It is helpful to get the bitch owner’s credit card information so that services and FedEx fees can be billed directly to them.

Same Day, Weekend and Holiday Shipments
1. Prior to 9/11 terrorist attack, counter-to-counter shipment of semen offered a convenient option for weekend inseminations. The dog owner transports the package to the airport where it is put on a flight (preferably nonstop) to the recipient’s nearest airport.
2. Since 9/11, counter-to-counter shipments, the sender must be a “known shipper” to be granted shipping privileges by airlines. Known shipper status involves contacting each airline directly,
completing necessary forms and arranging for on-site inspection. Delta, United and US Airways offer reliable services.
3. Fed Ex offers “Same Day Service” on a limited basis. The sender must be pre-qualified and inspected prior to using this service. Same day FedEx service is usually cost prohibitive.

Why – There’s only one good “why” – to make a better dog!


Summary
Great advances in canine assisted reproduction techniques have been achieved during the past 20 years. In addition to natural breeding, artificial inseminations with fresh “neat” (unextended) semen, fresh-chilled extended semen and frozen semen are now viable breeding options. With each technological advance, so to come more manipulations by human hands and an increased risk for human error in semen processing and insemination. Optimal results are best achieved when the breeding is carefully planned, ovulation is timed and a coordinated team effort is implemented.