What is sperm fragmentation assay?

DNA fragmentation test looks in detail into the DNA of the sperm. We know from research that highly fragmented DNA in the sperm can be a cause of infertility. So, when we look at the results and there are different ways, different labs do it differently but in general, any DNA fragmentation that is 15% or less, is excellent sperm so, we know that if we don’t have more than 15% DNA fragmentation, the sperm is healthy and should hopefully give a success result in IVF or even naturally, natural pregnancy. If it’s 15 to 25% fragmented then, it’s fair to good but anything more than 30% or 25 to 50%, we do say that is fair to poor. If more than 50% fragmentation then, a very big cause of concern because we know that the outcome of either the IVF treatment or even trying naturally will not be very successful.

DNA fragmentation tests can be done on a sperm sample. Man produces a sample, it goes off to an external lab and they look at the integrity of the DNA within that sample and they come back with a percentage of sperm which is damaged, degraded, highly degraded and it gives us different cutoffs to say actually this sample is good, it’s fine for a natural conception or for an IVF cycle or this DNA is more fragmented and you should be doing ICSI, even if the sperm parameters are normal just to try and minimize that DNA fragmentation or potentially PICSI or actually the DNA fragmentation is so high maybe the patient should consider using donor sperm.
The problem with it is that it only measures the DNA fragmentation in that one sample. Sperm production takes 81 days, if you do another sample three months down the line, the DNA fragmentation changed so it’s important to maybe do two if not three DNA fragmentation samples to build up a baseline like you would do with a normal sequence analysis, you wouldn’t just base a fertility assessment of one single analysis. It’s a building body of evidence suggesting that DNA fragmentation should be included as a routine diagnostic assessment and with every semens analysis but it can be quite expensive. IVFs are already quite expensive so adding on all these extra diagnostic tests is not in the patient’s best interest unless there’s a specific clinical history for it.

The Sperm Fragmentation Assay is very much a poor relation compared to DNA Sequencing and Karyotyping. No actual DNA letters are read in this assay. All the Sperm Fragmentation Assay detects is the number of unstable chromosomes present in a sperm population due to environmental factors like drinking and smoking or physiological factors like varicocele presence on the man’s testicles. Unfortunately, the correlation between observed increased fragmentation and failed infertility is poor. The only outcome of excessive fragmentation of ejaculated sperm is to take testicular sperm sample. However, this is not free of complications and should only be used as a last resort.

Sperm DNA fragmentation has emerged recently in the armamentarium of the evaluation of the male factor. Conventionally it used to be WHO semen analysis where we check the concentration, motility and normal shaped sperm. But clinically now, sperm DNA fragmentation tests are available, although different clinics complete them slightly differently and there are 2 or 3 essays that are available in the market with their own pro’s and con’s. The general consensus is that it is still not a mainstream must male factor test yet because of the limitations, although in certain cases it has advantages and we can assess the function of this sperms more closely, specifically is someone is modifying something which is modifiable in their case with low semen parameters and we can see if that has improved or not.
So, basically sperm is a complex cell and it contains a compact DNA that is affected by lifestyle, environmental toxins, diet and different other things and it can lead to fragmentation or breaks within the DNA. This has translated into either difficulty in conceiving for the female partner or there is now a role that if there is high DNA fragmentation or more DNA breaks in the sperm, then it leads to problematic integrity of the sperm and can translate into more chances of miscarriage because the embryo is 50 percent of the genetic component of the male as well. Previously we used to ignore males as well and simple semen analysis was sufficient for them and rule out abnormality or diagnose normality but now there is more and more focus on male health and the sperm micro structure and genetics. So now we have learned that yes, sperm that is damaged can lead to problems and we can clinically test this by doing DNA fragmentation tests with common essays being TUNEL assay or Comet assay. There are pros and cons for both of them but the main limitation is that the sperm sample that is used to check the DNA fragmentation is not the sample that is used on the day when we use it. Therefore it could be slightly different and we know that sperm parameters are very variable in a single man at different timings and different physiological conditions are under different lifestyle factors. So the problem is that the sperm sample that is tested for DNA fragmentation is not used in assisted conception, it is a different sample. So the reproducibility is also an issue and the standardisation of the lab assay is also problematic, it is not universal. That is why it has probably not been universally adopted by all the clinics. But obviously the clinics that have researched it and have standardised their assay’s have more robust meaningful reserves. It is an addition l in the diagnostic tests and to know more about the sperm health but it is not a must in my opinion.

Today, there is an agreement that DNA fragmentation tests of above 30% may indicate a degree of male infertility, so an additional degree of infertility. What we recommend for the men that have been diagnosed with increased DNA fragmentation, is to ask them to improve their lifestyle in an effort to improve the quality of the spermatozoa they produce. High DNA fragmentation may be identified in both oligospermic males and normospermic males – especially the ones that have been diagnosed with unexplained infertility. So with a DNA fragmentation that is rated above 30, a good way to go is to ask the patients first to improve their lifestyle, increase the intake of supplements, vitamins that have been shown that they contribute to reduce DNA fragmentation, increase the frequency of ejaculations, reduce the body mass in case there is increased BMI, stop smoking (quit smoking), increase sport activity, increase intercourse frequency in an effort to optimize the fertility and especially the quality of the spermatozoa they produce. Even if this male is oligospermic and he will eventually resort to a micromanipulation technique, even for this man, is crucial to improve the quality of the spermatozoa they produce because via ICSI, we do tend to select the good looking morphologically spermatozoa but we are not able in an efficient way to identify the spermatozoa through their morphology that have fragmented DNA. We do have a number of techniques that work as filters but again it is much better to start with a better sample from the very beginning and then implement all our tools and filters in an effort to further improve our results and the success rates. In cases however, where despite the efforts we do tend to get affected and above 30% DNA fragmentation, we may resort to testicular spermatozoa retrieval in an effort to have better quality spermatozoa and better DNA structure in them.

The sperm has got DNA in it like pretty much all cells and that’s what is taken to the egg, it’s in it’s injected into the egg at fertilization and it fuses with the eggs DNA to make an embryo. So DNA can be damaged by a number of things: damaged by for example radiation, chemicals etc. One of the ways in which the damage occurs is that they produce molecules within the cells that are harmful to the DNA. The damage to DNA as a result of perhaps heating or infection. Other things and that DNA damage can interfere with the sperm’s ability to develop a normal embryo, possibly to fertilize also. Tests for it look to see how much of the DNA in the sperms is fragmented compared to normal but there have been problems with these tests in terms of working out what normal should be for people and also from the point of view that if you go for ICSI, you would generally get a well-moving sperm that doesn’t actually perhaps have high levels of DNA fragmentation and you can’t necessarily pick out from a DNA fragmentation test an individual sperm or sperms that have low levels of DNA fragmentation. So it’s a bit limited in what it can do and that’s one of the reasons why they’re not really performed in NHS units.

Sperm chromatin fragmentation tests are tests to investigate if the DNA of spermatozoa has some damage. The most important tests to assess sperm DNA fragmentation are TUNEL or SCAS. SCAS is one of the most important techniques, it is carried out with flow cytometric and if the patient has more than 25% of spermatozoa.

We have to take into account that if we talk about DNA fragmentation, we are looking at the DNA of the sperm and which kind of breaks it has. There are different kinds of breaks: single strand break and double strand break. Depending on which DNA fragmentation we want to assess, we will do one test or another. There are many of them: Comet, SCD, Tunnel and so on. Depending on DNA fragmentation, we will have one diagnosis or another. We have to take into account that if we have high DNA fragmentation, it can be affecting the cycle by having lower blastocyst rate, lower implantation rate and higher miscarriage rate so usually with patients with this kind of background it is recommended to do this kind of tests. What can we do to improve DNA fragmentation test, usually it is recommended to take vitamins, change lifestyle but the most important is to visit a specialist, the Andrologist and they will recommend what is the best treatment for you.

It is an interesting question because the male factor is one of the 50% of reasons for infertility in the couple. The outcome of an IVF trial is to have a successful pregnancy and healthy embryo, the male factor is an important part.
One specific test is called DNA fragmentation and not many gynaecologists or many IVF centers support this test because what they look at is only for the specific sperm analysis.
Why is this inadequate for a successful IVF trial or even for a natural pregnancy? Because we have seen cases, especially the last 10 years, that the sperm analysis looked okay for some physicians while we see the morphology of the sperm analysis. If it’s below 10% for example, we’ve seen 2%, 4%, 5%, we see an immediate increase of the DNA fragmented material of the sperm. What does it mean? That means that half of the genetic material of the future embryo is not okay genetically. It means that even if we have a successful IVF pregnancy or natural pregnancy, we will see miscarriages at the first 3 or 4 weeks of the gestational – that’s why we have to be careful and ask for DNA fragmentation of the male sperm.

Sperm DNA fragmentation is related to any changes in the structure of the sperm and usually that involves a single or a double-strand break in the sperm. There are different tests that they examine the levels of fragmentation on the sperm DNA and those tests they need to be done carefully – obviously in the lab to give us accurate results and depending also on what sperm DNA fragmentation test has been done, those tests they might give us apart from the average score of the DNA fragmentation as the DNA fragmentation index as we may call, it they can show us that the level of the breaks we have on that DNA of the sperm and also the proportion – the average score, so it can you can show us the extent and also to the proportion of the sperm that they have the those fragmented DNA .
How can we use this information of what is the proportion of the sperm with their fragmented DNA and with high levels of fragmentation or sperm with low levels of fragmentation?
The clinicians can use that information to determine whether a patient would be benefited, for example, from the presence of any antioxidants and patients may take antioxidants for three months for example and then after those three months we’ll have to do another DNA fragmentation test to see if the integrity of the sperm DNA has improved or whether they need to do ,for example, testicular ultrasound to see if there is a varicocele going on which might also affect the quality of the DNA as a varicocele increases the blood flow within the testes, it can increase the temperature and that can increase the oxidative stress in the testicular environment and hence it can have an effect on the DNA integrity.
Also apart from the steps that can be followed prior to embarking on an a any ART techniques, if the couple has to use the assisted reproductive technologies by having a high sperm DNA fragmentation index, the fertility experts can decide what type of ART treatment can be followed so for example IVF versus ICSI or IMSI or PIMSI.

We had looked at lots of different aspects of sperm function: we looked at acrosome reactions, we looked at motility and we found that these weren’t relevant in every case. But no matter what we did for the difference between men who were having children and men who weren’t or people who had disease conditions like diabetes, there was always a difference in the quality of their sperm. Now the tests that we have, we call them the EXACT tests because they’re very accurate and we can give couples three different results from our EXACT test. The first one is the average of DNA damage across all the sperm that we measure in the semen, then we can tell them the population of sperm which are really good (which would be equivalent to those in a fertile man’s semen) and then, we can give them the population which are really bad. Now, all of those are really important because even the really bad ones usually have high reactive oxygen species or ROS and they’re the ones that can then damage the good sperm. So the more good sperm you have, the better – the less bad sperm you have, the better. So, those three results will then aid a couple both in diagnosing, if the man has a problem and secondly, in guiding him to whether IVF or ICSI would be the best treatment for him and also we have to remember that men unlike women are very fortunate in that they produce a new sperm every three months, so even if they get a bad result, they can make quite simple changes to their lifestyle like trying to lose a little bit of weight, stop smoking, not drink so much alcohol, don’t take recreational drugs and then in three months time their sperm quality could be much better than it had been. So it’s a very good news story and it’s something that a man can take control of his life and improve his sperm and make him more fit for fatherhood.

DNA fragmentation tests assess the presence of either single or double cell breaks in a sperm’s DNA. This is very important and considered an advanced test for the investigation for male infertility. It is important because high levels of sperm DNA fragmentation have been associated with a lower and worse reproductive outcome and the reduced male fertility potential. There are particularly specific groups of patients who can benefit from sperm DNA fragmentation testing. Different clinical societies have recommended that patients with unexplained or idiopathic infertility should be tested for sperm DNA fragmentation. Similarly, patients with clinical varicoceles or patients that are exposed to environmental factors that may impact male fertility or similarly important, couples who are experiencing recurrent pregnancy loss, it is important for them to be tested for sperm DNA fragmentation.
Why? Because if occurs that one couple has high sperm DNA fragmentation, there is some treatment that we can consider, for example, oral antioxidant therapy to modify lifestyle factors, if we identify some factors such as obesity, smoking or alcohol, impacting the rates of sperm DNA fragmentation. Similarly, control of infection and inflammation is also important as all of these conditions can increase the rate of fragmentation. The presence of clinical conditions should be treated. If you have varicoceles, it is important that you perform a varicocelectomy in order to reduce sperm DNA fragmentation. If a man keeps showing a high rate of sperm DNA fragmentation, it should be recommended to perform ICSI and not IVF in case of assisted reproduction. Similarly, sperm preparation techniques should be used in order to select 2 samples of sperm with lower sperm DNA fragmentation rate. Another possibility is to consider retrieving sperm directly from the testes instead of using sperm from the ejaculate because it has been shown that sperm from the testes has a lower rate of sperm DNA fragmentation than simply obtaining by masturbation from the ejaculation.