This workshop covered specific advice around bull handling, semen handling and evaluation techniques, providing tips on how to avoid artefacts in samples, so that veterinarians are able to trust their results.
Equipment set up
All measures should be taken to ensure that the sample seen down the microscope is a representative biopsy of what would be ejaculated during natural service. Cold shock is the most common and challenging hurdle to overcome, but preparation and use of suitable equipment can help avoid premature death and artificially low motility of cells. Lab equipment should be set up before the bull examination to allow the kit to reach the optimum temperature of approximately 37°C. For some kit it depends on what the manufacturer recommends.
Before arrival on farm, check there is a suitable electrical supply to set up all equipment. The lab area should be under cover and as close to the crush as possible.
Useful kit for conducting an examination includes:
- Small table top incubator to maintain temperature of equipment
- Slide warming bench
- Heated microscope stage
Pre-focusing the microscope is helpful. Keeping the collection tube warm is key.
Bull handling
Safety for handler and bull are paramount. A suitable crush and a suitable restraint is required, and a non-slip surface. A crush with a head yolk and a door in front of this is preferable, but they must not be able to duck out the side of the crush during collection.
Examination tips
The BCVA Bull Pre-Breeding Examination Certificate is referred to as the gold standard. When carrying out the scrotal circumference portion of the physical examination, hold the cranial portion of the neck of the scrotum, and place the measuring tape around the widest part below where you are holding. Do not hold the base of the scrotum.
After examining the internal sex glands, trans-rectal massage of the ampullae helps to pre-stimulate the bull before electo-ejaculator use.
There are no minimum standards when considering both the volume and appearance of the semen sample since it is not comparable to a natural collection, so it is not recorded during testing. However, it can be useful to note the appearance to estimate the concentration of the sample (Revel, 2010). This can help influence the size of the drop to be analysed and avoid overcrowding of the slide.
Sample collection
Avoid the probe losing contact with the rectal wall and sex glands. It is important to avoid sample contamination. Curved, blunt-ended scissors can help to clip hair around prepuce. Screw top lids on collection tubes are useful to ensure no rain water can get into the tube.
Morphological assessment
The workshop covered the traditional method of assessment using the electro ejaculator and microscope. Semen samples can also be submitted for laboratory computer assisted semen analysis and flow cytometry.
Assessment of motility is the first stage. Gross motility is scored from 1–5. A good semen sample should score at least 3.
Progressive motility is an assessment of the ability of the sperm to move through the reproductive tract. It is assessed by diluting the sample in pre-warmed phosphate buffer solution. The accepted minimum standard is 60% for BCVA, which is higher than other bodies who use 30%.
For morphological assessment it is important to avoid an overcrowded smear. Doing this bull-side can help avoid damage to cells.
A repeat sample should be taken from a bull with a progressive motility of less than 60%. With a continued motility failure you will likely see dead or abnormal cells. If inconsistent results occur this may indicate semen mishandling. Understanding normal sperm morphology is key so you can explain the findings to farmers and how these impact the breeding situation. There are at least 25 different sperm cell abnormalities. The following criteria can help with trying to establish the significance of the defect:
- What is the defect? Is it on the head, midpiece or tail?
- Where did it occur? In the teste, during maturation and storage, or during handling?
- The effect on fertility? Is it compensable or uncompensable?
It is important to have a good understanding of the process and what the findings mean because serious decisions are made on the basis of these examinations. Use the international standard of >70% of morphologically normal cells. In marginal cases, at least two 100-cell counts should be carried out. It is worth noting the live:dead count should also be carried out as well. It is reassuring that studies have found that decisions based on morphology investigations in the field, compared to those in labs who had better microscopes, were the same 92% of the time. Both beef and dairy bull morphology results are strongly correlated with fertility.
Satisfactory and unsatisfactory bulls are relatively easy to identify, but the marginal ones present a challenge. Where a bull passes all elements but fails on morphology, it is suggested to retest in 60 days time if possible. Young bulls (12–15 months) may have lots of abnormal looking semen cells – this can be known as an immature profile. Ideally wait 2–3 months and retest these individuals.
Conclusions
Preparation is key and taking the time to minimise adverse external factors is essential, so you are confident that what you see down the microscope is a real snapshot of the bull's fertility.