How Progressively Motile Are Those Sperm?
– A comparison of videos of CASA evaluations showing poor, good and excellent progressively motile sperm. A useful teaching aid for vets and breeders.
“Motility” evaluation – description and use.
A basic evaluation of a stallion’s semen will include a review of samples under the microscope to determine if and how the sperm are moving. This is typically achieved at a 100x or 400x magnification. Situations where such an evaluation would be performed are prior to shipment of semen cooled; prior to (or following) insemination of semen; or before, during, or after freezing of semen. There are two categories of motility – total and progressively motile sperm, which are usually expressed as a percentage of all sperm. “Total motility” indicates the overall percentage of sperm that are moving in any direction, whereas “progressive motility” indicates the percentage overall of sperm moving in more or less
a straight line. A further category of non-motile is applied to sperm that are not moving.
How straight is “straight”?
It is a generally accepted that for an equine sperm to be considered progressive, it should demonstrate 75% or greater straightness. To simplify: if a sperm were to travel 1 inch when going along in an averaged path of it’s actual convoluted track, a straight line drawn between the starting and end points would be no longer than ¾ of an inch long. With CASA (Computer Assisted Semen Analysis) a further figure is included in the equation, which is the speed of travel in those two parameters, so the full equation at which the percentage of “Straightness” (STR) is arrived measures the “Velocity Average Path” (VAP) and the “Velocity Straight
Line” (VSL) to produce the following formula:
Essentially therefore the actual “smoothed” path of the sperm can be no more than one-third greater than that of the straight line drawn between the start and finished points. This means that the difference between progressive or not can be subtle and therefore difficult to determine when reviewed by a human through a microscope – especially when one considers that these sperm are (hopefully) not lazily moving along, but are in fact quite rapid!
Subjective evaluation under the microscope.
The first point to make note of is that unless the raw semen sample is not particularly concentrated, it is almost impossible to reliable determine percentages of progressive/overall motility without dilution. Furthermore, some stallions have seminal plasma that is so toxic to sperm that unless the sample is diluted, the sperm will rapidly lose motility and die. Dilution with at least an equal part of extender by volume is therefore an important first step. It also will make it easier if the sample to be evaluated for motility is diluted to a concentration of 25 million sperm/ml or lower. In particular when first starting our evaluating semen, a lower sperm concentration is essential – ideally with around 20 sperm being visible on the individual microscope field of view. Note that while adding a smaller drop of semen to the microscope slide can achieve the lower number, if too small a drop is used, the lack of depth of liquid between the cover slip and slide may negatively impact sperm motility.
Objective evaluation using CASA
Computer-Assisted Semen Analysis takes the guesswork out of determining motility, be it progressive or total. The CASA unit takes a very short video of the semen (typically about one-half second) and compares the position of all the sperm in each frame during that short time period. A computer then evaluates the differences in the sperm movement and provides a variety of different results including total and progressive motility. When one uses CASA, it quickly becomes apparent that for many people, their estimates – particularly of progressive motility – under the microscope is an over-estimate. One frequently hears stallion owners commenting “my stallion has excellent motility – he’s 70% progressive at 24 hours”. In fact, it is rare that one sees 70% progressive motility immediately after collection, never mind at 24 hours! Typically 50-60% progressive motility can be considered to be good – above that is exceptional!
Motility videos using CASA
Below we have a variety of videos of CASA evaluations. These have been recorded using one of our Hamilton-Thorne CEROS I Computer Assisted Semen Analysers. Each video starts by showing the sperm moving, then you will see the button being clicked to activate the CASA evaluation, then the results will show. After that on some of the videos, you will see a screen where you can view each sperm track. Different colours of track mean different things in those images: the blue/turquoise tracks are considered “progressive”; the green tracks are merely “motile” (i.e. not progressive); while the sperm that have a little red dot on them are considered non-motile. We would encourage you to start each video and determine your own estimate – pausing it before it reaches the CASA reading on order not to cheat (we give you about a 5 second warning)! If you need a little longer to get your own personal estimate, “rewind” the video back to the beginning and start again. We think you may well be surprised at the difference between your estimate and the actual CASA readings!
(You can go “full screen” by clicking the screen icon in the bottom right corner of each video when playing. )
Average Progressively Motile Sperm
Object lesson: It’s not the percentage progressive motility that is important; it’s the number of sperm that are progressively motile!!
Good Progressively Motile Sperm
Object lesson: You cannot “guesstimate” concentration just by looking at a sample under the microscope!!!
Object lesson: You must dilute samples prior to evaluation of motility. Ideally the concentration once diluted will be 25 million sperm/ml or less
Poor Progressively Motile Sperm
Object lesson: Progressive motility and velocity are important – but remember that those figures need to be applied to total sperm numbers to get a meaningful statistic!
Numbers not percentages!
|We cannot stress this enough! Many people get “hung up” on percentage progressive motility and voice major concerns when they receive semen that is say 10% progressive. Well, of course one would prefer to receive semen with sperm that are more progressive than that, but without looking at the actual numbers of sperm involved, one is not seeing the whole picture. This is very clearly explained by looking at the following sets of figures:|
|% Progressive Motility||Number of sperm in shipment||Number of progressively motile sperm in shipment|
|60||250 million||150 million|
|10||2 billion||200 million|
|So as you can see from the above numbers, in fact the sample with 60% progressive motility doesn’t have as many progressively motile sperm as the sample with 10% progressive motility!
Object lesson: One should not concentrate solely on the percentages!
Key points To Consider
- Be tough on your visual evaluations of sperm motility – in particular progressive
motility! Remember that 60%+ progression is a rare entity in real
- Do not attempt to determine motility levels with too concentrated a sample – it
can’t be done with repetitive accuracy!
- You cannot “guesstimate” sperm concentration using a plain microscope slide. If
you are going to count it under a microscope, use a hemacytometer or counting chamber
of some sort. Alternatively obtain an estimate using a mechanical sperm counter.
- Don’t worry too much about the percentages – worry about the numbers! Multiply
percentage motility by sperm numbers present to obtain the actual number of
progressively motile sperm. That is a significant statistic.
- Pay attention to insemination dose numbers, but don’t rely on old research!
>100 million progressively motile sperm is enough to do the job presuming close
proximity of insemination to ovulation.
- Remember that progressive (or any) motility is not an absolute guarantee of
fertility! While it’s a good start, it is still possible to have good progressive
motility, poor or even absolutely zero fertility! This is particularly the case with
frozen semen, where the cell may be negatively affected during the freezing/thawing
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