Semen Evaluation Flashcards
indications of semen evaluation (8)
- Breeding Soundness Certification.
- Before breeding/purchase for breeding
- Diagnosis and/or prognosis of reproductive disorders.
- Characterize semen samples for trade.
- Processing insemination doses (fresh/ chilled/ frozen semen).
- Guide clinical and management decisions.
- Assessment of treatment / sperm production (e.g., toxicology and nutrition studies).
- Frozen straw quality control
semen collection with an artificial vagina; stimulates, species used in, pros, cons
- Simulates natural mating (shaping, texture, warmness, slipperiness, attitude).
- Commonly in bulls, stallions, rams, and goats.
Pros
* Comfortable for male, yields physiological semen
Cons
* Requires training to mount dummy/teaser,
* Not suitable for all species or individuals (uncooperative or physical limitations)
semen collection with an electroejaculation; stimulates, species used in, pros, cons
- Probe into the male rectum,
- Delivers mild electrical stimuli to induce ejaculation.
- Commonly in bulls, rams, and sometimes in wildlife species.
Pros
* Does not require prior training,
* Useful for collecting males that are unable or unwilling.
Cons
* May cause discomfort or stress to the animal.
* Semen quality sometimes lower compared to other methods
semen collection with an manual massage; stimulates, species used in, pros, cons
- Massages the animal’s penis to induce ejaculation.
- Commonly in Dog, Boar, Poultry.
Pros
* Does not require specialized equipment.
* Can be performed without the need for a teaser animal.
Cons
* Requires skill and experience
* May be time-consuming
what are sperm cells highly sensitive to? (7)
- Temperature: sudden changes, abnormal hot/cold
- pH: Acidic or alkaline environments damage sperm cells.
- Osmotic Pressure:Hypotonic or hypertonic cause swelling or shrinkage
- Light Exposure: Prolonged exposure to light, especially UV light.
- Toxic Substances: Detergents, disinfectants, and heavy metals
- Time: sperm loses viability over time.
- Mechanical Stress: vigorous agitation
things to do with semen handling (9)
- Use clean, non-reactive materials (e.g., AVs, tubes, syringes, slides, pipettes).
- Pre-warm all tools and equipment (~37–38°C).
- Avoid sudden temperature changes.
- Handle semen gently.
- Keep samples away from light and air.
- Avoid contact with water, detergents, or chemicals.
- Train staff on correct techniques.
- Update lab protocols regularly.
- Record details for every animal and sample
macroscopic evaluation of semen (4)
o Volume
o Colour
o Consistency
o pH
routine evaluation of semen (5)
o Concentration
o Motility
o Morphology
o Viability
o Culture
advanced evaluation of semen (4)
o DNA Fragmentation
o Mitochondrial Activity
o Oocyte penetration
o Proteomic and Genomic
features of volume evaluation and influence
- Stallions (gel-free) 30–100 ml, Bulls: 2–10 ml, Dog (sperm-rich) 0.5–3 ml.
- Influence of species, breed, individual, method & frequency of collection, teasing, season
normal vs abnormal colours of semen
- Normal: Milky white to creamy
- Abnormal Colors:
o Pink/Red: Hemospermia (trauma/ inflammation reproductive tract)
o Yellow: Urospermia, indicating urine contamination.
o Greenish: Suggests pyospermia (infection).
o Translucent: Low sperm concentration
normal vs abnormal consistency of semen
Normal
* Semen appears slightly viscous and turbid, with a uniform texture.
Abnormal
* Watery: low sperm conc., excessive collection frequency , testicular hypofunction.
* Thick or Gel-Like: stallions excess seminal plasma proteins or prolonged abstinence.
normal vs abnormal pH of semen
Normal
* Stallions: 7.2–7.8…..Bulls: 6.8–7.2
* Dog: 6.5–7.0 for the sperm-rich fraction.
Abnormal pH
* Acidic: Urospermia or inflammation
* Alkaline: Infections or contamination
what is a hemocytometer used for
are the most common, the use of chambers in which cells can be
observed and counted within a known area, then allowing the calculation of the number of
sperm per unit of volume
pros and cons of hemocytometers
Pros: Low cost and widely accessible. Direct visualization allows confirmation of sperm morphology and exclusion of debris.
Cons: Labor-intensive and time-consuming. Requires skilled technicians to ensure accuracy.
Errors can arise from improper dilution, uneven sample mixing, or misinterpretation of grid boundaries
what is a Spectrophotometer used for
estimate sperm conc. based on the optical density or turbidity of a semen sample. Sperm conc. is directly proportional to its optical density when measured at a specific wavelength of light. As sperm cells scatter light, the higher the sperm concentration, the greater the light scattering
pros and cons of Spectrophotometers
Pros
* Quick and efficient for high sample volumes.
* Relatively inexpensive equipment.
* Minimal sample handling compared to manual methods.
Cons
* Indirect measurement can be influenced by non-sperm (debris or other cells).
* Requires calibration for each species
* Suitable for raw but not extended semen (extender interfere light)
what is a nucleocounter used for
a fluorescent dye (propidium iodide) to stain sperm DNA. Measures
fluorescence intensity to count sperm within a defined volume. First, dilute semen sample with detergent that damage the sperm membrane and allow PI to bind DNS
pros and cons of a nucleocounter
Pros
* Highly accurate and specific to sperm, excluding non-sperm components.
* No calibration required, as sample preparation follows a standardized protocol.
* Quick analysis (approximately 30 seconds per sample).
Cons
* Requires proprietary consumables (diluents and disposable cassettes).
* Influence of somatic cells (contains DNA, ex: WBC, Epithelial cells)
* Higher cost compared to hemocytometers.
* Limited to species pre-validated by the manufacturer
how to evaluate gross motility of sperm
- Single drop of undiluted semen + no coverslip is evaluated at 10×.
- Affected by the concentration of the sample.
- Swirling, Waves, Eddies – only in species with high sperm conc. (bull, ram, buck)
motility; normal, motile vs non motile sperm for fertilization
- Historically, the most common test of sperm quality (easy to-do)
- Normal: bull 30-60% stallion 60%, Dog 70%
- Non-motile sperm definitely unable to Fertilize the ova.
- Motile sperm may/may not be able to Fertilize the ova (Motility poorly correlated
with fertility, but still important parameter). - Many morphologic abnormalities are motile!
motility is a function of _____ and ____
midpiece and tail
gross motility scores (4)
Very Good (VG): rapid dark swirls and eddies
Good (Good): slower swirls and eddies
Fair (F): no swirls, but prominent individual motion
Poor (P): little or no individual cell motion
what is individual motility influenced by
Morphology, temperature, time between collection & evaluation, detergents, lubricants, latex, water, contamination with dirt, epithelial cells
two main aspects to assess for individual motility
total motility, progressive motility
visual inspection of sperm and classification
- Subjective assessment of sperm motility
- Phase-contrast microscopy
- Estimate if 50, 75, 25% of the sperm that are in motion
what is computer assisted semen analysis used for
- use automated software combined with microscopy and video imaging to analyze parameters such as sperm motility (progressive and total), velocity, concentration,
morphology, and other kinematic traits (straightness, and linearity) - Useful in research arena . objectivity and repeatability measurements.
what is the most useful and important aspect of sperm quality? methods
- Sperm morphology is the most useful and important aspect of the
semen quality Not time dependent, can fixed evaluated at a later - The eosin–nigrosin (EN) staining For (Morphology+ Viability)
- Eosin is a vital stain penetrates dead sperm (pink) ,
- Nigrosin provides a dark background for contrast
__% normal sperm is the threshold for a satisfactory sample
70%
categories of sperm morphology
Each sperm is placed into 1 category, only: Normal or Abnormal head, midpiece, and tail defects
types of abnormal sperm morphology (9)
A) Normal sperm
B) macro, micro, tapered, pyriformhead , constricted, degenerate head
C) Acrosomal defect
D) Proximal and distal cytoplasmic droplets
E) segmental aplasia, rough, enlarged mitochondrial sheath, bent midpiece, double midpiece/double head
F) Bent tail (or hairpin tail),
G) Coiled tail, with/ without encircling the head
I) Premature germ cells with a single /multiple nuclei
what is the dag defect
characterized by a figure -8- appearance of the midpiece. caused by gossypol toxicosis
features to become a satisfactory potential breeder (4)
-Passed a physical exam.
-Met the minimum requirements for scrotal circumference based on his age/breed
-Has at least 30% (SFT) or (60%) WCABP progressive sperm motility.
-Produces at least 70 % normal sperm
