Page 556 - Equine Clinical Medicine, Surgery and Reproduction, 2nd Edition

Reproductive system: 2.2 The male reproductive tr act 531

VetBooks.ir aberration in an animal with a suspected intersex without contamination of cells from other sources.
Live lymphocytes are cultured for 72 hours and the
condition.
Aberrations of the autosomes are rare, as they
generally lead to embryonic failure early in develop- nuclear chromatin collected and stained to perform
the karyotype analysis. Specific C- and G-banding
ment. Sex chromosome abnormalities are the most techniques are used to identify specific chromo-
common finding. Two sodium heparinised tubes of somes. Fluorescent in situ hybridisation and PCR
freshly collected blood are submitted on ice over- techniques are used to probe for translocations,
night to cytogenetic laboratories for testing. The duplication and deletions, including SRY expression.
blood sample should be collected aseptically and

ADVANCED REPRODUCTIVE TECHNIQUES

CHILLED SEMEN pursues a performance career or is sidelined by injury
or disease. Cryopreserved semen appears to retain
The transport of chilled extended semen is now its fertilising ability virtually permanently, provided
commonplace and has greatly influenced the growth it is stored properly in liquid nitrogen. The semen
of AI. Transport of equine semen across state lines, from individual stallions varies considerably in
countries or continents may require that govern- response to cryopreservation. While most stallions’
ment permits, certification of disease-free status or frozen semen will result in acceptable fertility rates,
other paperwork is completed. Many commercially the semen from some individuals cannot be frozen
available transport packaging systems are now avail- successfully. The reasons for individual variation in
able, ranging from short-term (24 hour) cardboard response to semen cryopreservation are not known.
boxes with icepacks to the longer-term (48–72 hour) A stallion with poor-quality semen is unlikely to
Equitainer. Research has shown that most stallions’ be successful in a cryopreservation programme.
semen retains best motility when extended for chill- Perhaps more frustrating is the stallion with high-
ing to a concentration of 25–50 million sperm/ml. quality semen and good post-thaw motility that
Following routine semen collection and analysis, achieves few or no pregnancies with cryopreserved
semen is extended and packaged in a leak-proof con- semen. Per cycle pregnancy rates are generally lower
tainer such as a Whirl-Pak bag. Clear labelling of the than that expected with natural cover, fresh or trans-
bag and the exterior of the container should include ported breeding programmes. Several studies have
the following information: stallion name; owner; demonstrated that average per cycle pregnancy rates
registration number; collection date; motility, con- with frozen equine semen are in the 50–70% range
centration and number of sperm shipped; and mare when mares are intensively monitored and insemi-
identification. It is generally accepted that 500 mil- nated very close to ovulation. Practitioners should
lion motile sperm should be included in an insemina- discuss the expected results with clients wishing to
tion dose, although some stallions may demonstrate use frozen semen in their breeding programme.
acceptable fertility with as low as 200 million motile The cryopreservation process involves routine
sperm per dose. If semen will not reach the mare for semen collection, centrifugation to remove the
24–48 hours, the expected decline in motility should seminal plasma, resuspension in freezing extender,
be factored in and a higher number of total sperm packaging, cooling and, finally, freezing in liquid
included in the shipment. nitrogen. Depending on the stallion and extender
used, semen may or may not be chilled prior to cryo-
SEMEN FREEZING preservation. The availability of commercial, ready-
to-use extenders for cryopreservation of stallion
Cryopreservation allows permanent storage of sperm has greatly simplified semen preservation.
semen, worldwide distribution of superior genet- Freezing extenders contain buffers, antibiotics, sug-
ics and permits continued breeding while a stallion ars, egg yolk and a cryoprotectant such as glycerol,