Contributors: Vetstream Ltd
Species: Canine | Classification: Lab Tests
- Examination of semen sample provides information about breeding problems in stud dog.
- Should be performed in conjunction with full breeding health assessment Breeding soundness examination.
- This technique is multifaceted and can stand alone.
- To investigate poor fertility in stud dogs.
- Palpation of prostate gland + ultrasonography of gland in older dogs and those with urethral discharges. Rectal examination of prostate important to aid in assessment of size and consistency.
Since rectal examination may cause discomfort it is best delayed until after semen collection.
- Visual examination of preputium/penis in erect/non-erect states.
Source of Test Material
- Testes, epydidymus and prostate gland.
Quantity of Test Material
- 0.5-1.5 normal range (smaller quantities for smaller breeds. Larger amount if entire post-sperm fraction collected).
- Canine semen has 3 fractions:
- Pre-sperm: small volume clear fluid ejaculated as dog becomes sexually aroused and attempts, with thrusting movements, to achieve intromission.
- Sperm-rich: white color, ejaculated when 'tie' between dog and bitch commences, usually just before dog turns to face backwards away from bitch. No thrusting movements.
- Post-sperm: clear, voluminous, prostatic fluid which is ejaculated for several minutes during 'tie'. May be pause <2 minutes between ejaculation sperm-rich and post-sperm fraction.
- Volume of semen collected dependant on quantity post-sperm fraction included. Very small quantities with minimal post-sperm fraction may reduce likelihood of fertilisation.
- Sperm output: total number of sperm produced in ejaculate better parameter than sperm count or semen volume. 100 million normal live sperm in ejaculate is ideal target.
Sample Collection Technique
- Digital masturbation to produce semen sample Semen collection.
Take care not to damage penis with collecting vessel as may → hemorrhage → toxic effects on sperm.Do not use rubber latex for collection as has toxic effect on sperm.
- Examination of the motility, morphology and total numbers of spermatazoa.
- Palpation and ultrasonography of scrotum + contents.
Timing of test
- Pre-breeding, especially if using shipped, cooled semen.
- In cases of infertility.
- Semen samples should be evaluated immediately following collection.
- Cold shock can suppress motility and change morphology, eg folding of sperm tails.
- Classification of sperm into different types is not necessary.
- There should be no, or only occasional, inflammatory cells and no red blood cells in semen sample.
- Widely available.
- Probably highly specific.
- Probably low (+ve/-ve predictive values not calculated).
- Ratings, eg satisfactory, questionable, unsatisfactory, based on potential breeding capacity.
Technique (Intrinsic) Limitations
- Good +ve predictive value.
Only gross abnormalities detectable with this test.
Technician (Extrinsic) Limitations
- Level of skill dependant on familiarity of operator with normal anatomy and spermatology.
Normal (Reference) Values
- Characteristics of normal semen:
- Varies depending on post-sperm fraction included, eg normal: 5-15 ml; toy breeds: <5 ml; greyhound; >25 ml.
- Place drop of semen on warm slide (37 - 39°C) and examine under low power.
- The motility is expressed on a scale of 0-5, eg 0=nil motility; 5=very active sperm which may move in wave motion when shoals of sperm move around slide; optimum motility=3.
- Progressive motility:
- Place drop of sperm on warm (37-39°C) slide, under cover slip and examine under high magnification.
- Observe 100 sperm at random, the number moving forward in a straight line are counted.
- This gives a % figure.
- The optimum progressive motility= >70%.
- Sperm count:
- Mix 3 seminal fractions to ensure equal distribution of sperm.
- Use formal saline as a diluent and a red cell hemocytometer to count number of sperm in sample; 10,000-20,000 sperm/cu mm normal.
- The volume of the ejaculate is also measured and the sperm output calculated using the formula: sperm output = sperm count/cu mm x 1000 x sperm volume (ml).
- % live + normal sperm:
- After mixing 6 drops semen with nigrosin eosin stain leave for 5 minutes then make smear in same manner as blood smear.
- Dead sperm appear purple, live sperm do not take up stain.
- The smear is examine under an oil immersion lens and 100 sperm are counted to determine, eg % live/% dead sperm (target >70% live); % normal morphology (target >75% normal); % live and normal (target >70% live and normal).
Errors and Artifacts
- See artifacts.
- Incomplete ejaculate due to semen collection procedure (check alkaline phosphatase concentration).
- High proportion of bent tails and poor motility due to cold shock of semen or osmotic imbalance of stain (detached heads are seen in some cases of testicular degeneration). High numbers degenerate neutrophils and bacteria due to preputial content in semen sample.
- Large proportion dead sperm due to uses of some types of rubber in semen collection equipment.