Looks Can Be Deceiving: Advanced Semen Testing Can Predict Male Factor Infertility

 

Many of our new male fertility patients are perplexed about their inability to conceive. They have been told that it is not a female factor and that their semen quality is excellent. Their sperm have perfectly shaped oval heads, normal-sized mid-pieces, and straight, well-aligned tails, and they demonstrate great motility (i.e., sperm movement) and forward progression. On visual inspection, one would assume that their sperm could fertilize an egg with ease producing a normal embryo that would develop into a beautiful baby.

Unfortunately, in spite of their good looks, their sperm may never be baby-makers. The integrity of sperm DNA might be impaired. Even if sperm is able to fertilize an egg, DNA fragmentation will most likely prevent the normal development of an embryo, resulting in a failed pregnancy or miscarriage. I last wrote about sperm DNA fragmentation six years ago. New assays have been developed, and the literature updated.

It’s a growing concern amongst fertility specialists that sperm DNA fragmentation might be the cause of many couples’ inability to achieve a pregnancy naturally or through an assisted reproductive technology protocol using intrauterine insemination (IUI) or in-vitro fertilization (IVF) with or without the use of intracytoplasmic sperm injection (ICSI). I have added sperm DNA fragmentation testing to the battery of tests we perform to evaluate a man’s fertility potential. It can detect a reason for the couple’s subfertility and offer additional treatment options. We routinely screen our male patients with advanced male age, large varicoceles presence, or a history of miscarriages. 

We now also screen the male partner of infertile couples with a normal semen analysis and no obvious female factors for DNA fragmentation. In doing this, we have found greater than 40% of these men have abnormal levels of fragmentation. For these patients, this has answered questions they had and has offered them a better understanding of why certain options might be better for them in their quest towards parenthood. I will discuss some of the biology and therapeutic options in this blog. We have provided links for those wanting additional information.

DNA is an organic molecule that consists of two strands of repeating building blocks called nucleotides. Each nucleotide consists of a sugar, a phosphate group, and a nitrogen base. The nucleotides in each strand are tightly bonded. The two strands however, are loosely held together by weaker bonds. DNA fragmentation occurs when separations and breaks occur in these bonds. Our chromosomes are composed primarily of DNA, and segments of that DNA form genes, whose specific sequence of nitrogen bases constitutes genetic code. DNA fragmentation can impair the transmission of this genetic code, disrupting the cell’s ability to function. For a sperm cell, DNA fragmentation can prevent fertilization or proper development of an embryo if fertilization occurs.

https://www.halotechdna.com

Scientists have identified several factors that contribute to an increase in sperm DNA fragmentation. Oxidative stress is a major cause of DNA damage. Oxidative stress is the damage caused by free radicals reacting with molecules like DNA and thus disrupting their bonds. Physical and environmental changes can increase the level of free radicals to which sperm DNA is exposed. Recent studies have linked environmental toxins in certain chemical air pollutants, pesticides, and plastics to elevated oxidative stress and sperm DNA fragmentation. Obesity, alcohol consumption, and tobacco have also been associated with DNA damage from increased oxidative stress. 

Scrotal varicoceles have been shown to exacerbate sperm DNA fragmentation. Varicoceles elevate the temperature inside the testes to levels that disrupt the bonds of DNA molecules, resulting in fragmentation. Excess fat around the scrotum from being overweight will also elevate testicular temperatures to unfavorable levels that damage the DNA bonds.

Chemotherapy and radiation therapy have been linked to sperm DNA fragmentation, and further investigation is needed to determine if this is a long-term or even permanent side-effect of some cancer treatments. Reproductive aged men who have been diagnosed with cancer should be encouraged to cryopreserve sperm prior to starting treatment as a means of preserving their fertility. Post-treatment fertility evaluations should include an assessment of sperm DNA fragmentation, especially since men with normal semen analyses can have high levels of sperm DNA fragmentation.

Anomalies in the process of sperm maturation can cause sperm DNA fragmentation. The structure of the human sperm cell makes it more susceptible to DNA fragmentation than any other cell in the body. Its nucleus has 40% less space compared to most other cells, so sperm DNA has to be highly compacted and coiled to enable it to fit in the nucleus. This highly compacted and structured arrangement protects the sperm cell’s genetic material (DNA) as it travels through the male and female reproductive tracts, and it enables the proper fusion of DNA from both sperm and egg at fertilization. Sperm maturation is a process that transforms a round germ cell into an elongated mature sperm cell with a head, acrosome, midpiece, and a tail. During this process, DNA is uncoiled, transcribed (which is the process of reading the DNA’s genetic code), and recoiled a couple of times. Intentional nicks and breaks are made in sperm DNA to enable it to be tightly compacted and recoiled. These nicks are supposed to be repaired when the DNA is being transcribed, but external factors, such as oxidative stress and advanced paternal age, often prevent this and hence result in sperm DNA fragmentation.

Sertoli cells which function to support and nourish developing sperm cells in the testes also screen sperm cells and mark defective ones for elimination. Sometimes external factors disrupt this process, and marked sperm cells are not eliminated, or apoptosis (cell death) is initiated but not completed. These sperm cells have a much higher incidence of DNA fragmentation.

Sperm DNA fragmentation can occur during the movement of sperm from the seminiferous tubules through the vas deferens, an event that allows immature and mature sperm to be tightly packed together. Immature sperm produce a high level of reactive oxygen species (a type of free radical), which often react with and damage the DNA of mature sperm. Reactive oxygen species also activate enzymes called caspases and endonucleases that are part of the process of apoptosis, further damaging sperm DNA. This is the primary reason that sperm retrieved from the testes has less DNA damage than sperm from the ejaculate.  

Men with high levels of DNA fragmentation now have options. For many men, simple lifestyle changes can improve sperm DNA fragmentation. Weight loss, healthy diet, cessation of smoking, reduced consumption of alcohol, and antioxidants have all been shown to improve DNA integrity. For older men, who typically experience DNA damage because the mechanisms of sperm maturation function less efficiently, and men who have undergone treatment for cancer, it may not be possible to reduce sperm DNA damage. However, testis sperm have less sperm with fragmented DNA. Therefore, sperm can be surgically retrieved from the testis and used in IVF/ICSI. For additional information on sperm DNA fragmentation testing please contact me at bgilbert@nycryo.com.

References: 

 Ribas-Maynou, J., García-Peiró, A., Fernández-Encinas, A., Abad, C., Amengual, M., Prada, E., Navarro, J., Benet, J. (2013). Comprehensive analysis of sperm DNA fragmentation by five different assays: TUNEL assay, SCSA, SCD test and alkaline and neutral Comet assay Andrology 1(5), 715 722. https://dx.doi.org/10.1111/j.2047-2927.2013.00111.x

 Smit, M., Romijn, J., Wildhagen, M., Veldhoven, J., Weber, R., Dohle, G. (2013). Decreased Sperm DNA Fragmentation After Surgical Varicocelectomy is Associated With Increased Pregnancy Rate Journal of Urology 189(1S), S146-50. https://dx.doi.org/10.1016/j.juro.2012.11.024

 Tirado, E., Barrett, B., Leader, B. (2012). Concurrent sperm DNA fragmentation and oxidative stress assessment on 2,281 male semen samples Fertility and Sterility https://dx.doi.org/10.2164/jandrol.110.010595/full

 Bungum, M. (2012). Sperm DNA integrity assessment: a new tool in diagnosis and treatment of fertility. Obstetrics and gynecology international 2012(4), 531042 6. https://dx.doi.org/10.1155/2012/531042

 Moskovtsev, S., Mullen, J., Lecker, I., Jarvi, K., White, J., Roberts, M., Lo, K. (2010). Frequency and severity of sperm DNA damage in patients with confirmed cases of male infertility of different aetiologies Reproductive BioMedicine Online 20(6), 759 763. https://dx.doi.org/10.1016/j.rbmo.2010.03.002

 

Update #1: SARS-CoV-2 in Semen – Should we be concerned?

With the recent report that SARS-CoV-2 is found in semen, the answer now is; We probably should be concerned. This Research Letter published in JAMA Network Open by Li et al. (1) describes their findings in 38 participants who provided a semen specimen. They found

“23 participants (60.5%) had achieved clinical recovery, and 15 participants (39.5%) were at the acute stage of infection. Results of semen testing found that six patients (15.8%) had results positive for SARS-CoV-2, including 4 of 15 patients (26.7%) who were at the acute stage of infection and 2 of 23 patients (8.7%) who were recovering.”

The study, unfortunately, has several limitations. Thirty-eight patients are a small number, and there was no follow up to see when the virus that was found in semen was cleared from semen. Also, the two “recovering” patients were defined as patients that had a “clinical improvement,” which was not defined. They also did not measure the presence of the virus in the nasopharyngeal region at the same time as they test the semen. Other studies have found that SARS-CoV-2 can be found in oropharyngeal secretions for a mean of 9.5 days with a range of 2 to 22 days after symptoms began (2). This same study showed that the virus persisted in stool samples for 2 to 3 days longer than oropharyngeal secretions. Therefore, the two days that Li et al. study followed patients after symptoms started to improve was likely too short. In addition, studies are finding that as many as 20% of patients with the virus are asymptomatic and might be able to transmit the virus for a month or more (3). Therefore, a longitudinal study, following COVID-19 patients over time, measuring the time for viral clearance in several sites, including semen, is needed.

The results of this study also are in contrast to prior studies. I reported in two prior blogs that investigators did not find SARS-CoV-2 in semen or in testicular tissue (4) (NYCryo.com/blog). There is also a recent case report that also did not find SARS-CoV-2 in semen in a COVID-19 positive patient (5). All tests to date have used nasopharyngeal or oropharyngeal swabs to test semen specimens. However, not all body fluids are alike. They contain different proteins that might change the testing results. That is why a viral test must be validated for the body fluid it is being measured in. Validation minimizes the number of false positives (i.e., the test says the patient has the virus but actually the patient is free of the virus) and false negatives (i.e., the test says the patient does not have the virus but actually the patient has the virus). The lack of using validated tests is a possible reason why prior studies did not find the virus in semen (false negative). Conversely, it might also be the reason the Li et al. study found the virus in semen when it might not have been (false positive). To give valid results, the test must be validated for both the virus and the body fluid in which the virus is suspected to be residing.

We still don’t know if the virus in semen can be transmitted to the partner and cause disease. However, given the likely presence of the virus in semen, caution is advised for couples having relations. Especially when the male partner is positive for SARS-CoV-2. In an abundance of caution, until we have more information, abstinence or a barrier contraceptive should be recommended. 

I again refer to an editorial by Feldmann in the New England Journal of Medicine who offers a modified view (6). Dr. Feldmann made a point of differentiating between the presence of a virus detected in a semen sample and the infectivity of that virus. Many viruses are found in semen. However, they are not infectious. A measurement of infectivity is needed. To measure infectivity, you need to know the amount of virus required to cause an infection as well as the pathway that results in infection. For example, if a pathogen such as a virus is in semen but needs to be aerosolized and inhaled to become infectious, then it is less likely to be transmitted to another person. However, other pathogens such as Zika and Ebola can be transmitted by semen to another person through sexual contact. It appears, from the limited information available, that SARS-COV-2 is transmitted through transfer into the lungs by the act of breathing in aerosolized particles containing the virus or transfer of the virus from a surface containing the virus to a person’s mouth or nose even from COVID-19 positive individuals without symptoms (7). Hence, the requirement for hand-hygiene and social distancing. However, Dr. Feldmann feels, it is unlikely that sex, even if the virus is in semen, would be a significant modality of transmission. 

We also don’t know whether SARS-CoV-2 can be transmitted to the developing fetus. A recent case report (8) presented a second-trimester miscarriage in a suspected COBID-19 positive mother. The mother, placenta, and fetus were tested for SARS-CoV-2. The mother and placenta were positive, while the fetus was negative. The suspected cause of the miscarriage was SARS-CoV-2. It is difficult to make broad conclusions from this single case report. However, this report does raise additional concerns and challenges for pregnancies during this pandemic.

As I mentioned in a prior blog, the Society for Assisted Reproductive Technologies (SART) has recommended during this pandemic to stop new assisted reproductive procedures except for urgent cryopreservation (9). This is still good advice in light of the morbidity and potential mortality of this disease to the mother and child, as well as the poorly defined modality of transmission. 

My closing thoughts are updated based on this new information: SARS-CoV-2 are likely found in semen, they can survive the freeze/thaw process of cryopreservation and potentially can be transmitted to the partner and offspring. However, there is presently no data suggesting that SARS-CoV-2 can be transmitted to the partner and offspring from either fresh or cryopreserved semen. Pregnant women and their unborn child are a vulnerable population that should take all precautions for preventing SARS-CoV-2 infection during their pregnancy. Couples considering starting a family during this pandemic might consider being tested for the presence of the virus to make a more informed decision. Cryopreservation of semen should continue during this time with testing the patient for the presence of SARS-CoV-2 and if positive, quarantining the specimens with cryopreserved aliquots available for viral testing once testing of semen and testicular specimens is validated. Patients, as always, should be made aware of the limited data available and that the specimens should be quarantined until validated testing is available, and high-quality evidence regarding the use of SARS-CoV-2 positive specimens is available.

#covid19 #infertility #semen #coronavirus

References:

  1. Li, D., Jin, M., Bao, P., Zhao, W., Zhang, S. (2020). Clinical Characteristics and Results of Semen Tests Among Men With Coronavirus Disease 2019 JAMA Network Open 3(5), 1-3
  2. Ling, Y., Xu, S., Lin, Y., Tian, D., Zhu, Z., Dai, F., Wu, F., Song, Z., Huang, W., Chen, J., Hu, B., Wang, S., Mao, E., Zhu, L., Zhang, W., Lu, H. (2020). Persistence and clearance of viral RNA in 2019 novel coronavirus disease rehabilitation patients Chinese Medical Journal 133(9), 1039-1043. https://dx.doi.org/10.1097/cm9.0000000000000774
  3. Pan, Y., Yu, X., Du, X., Li, Q., Li, X., Qin, T., Wang, M., Jiang, M., Li, J., Li, W., Zhang, Q., Xu, Z., Zhang, L. (2020). Epidemiological and clinical characteristics of 26 asymptomatic SARS-CoV-2 carriers. The Journal of infectious diseases
  4. Song, C., Wang, Y., Li, W., Hu, B., Chen, G., Xia, P., Wang, W., Li, C., Sha, J., hu, z., Yang, X., Yao, B., Liu, Y.(2020). Detection of 2019 novel coronavirus in semen and testicular biopsy specimen of COVID-19 patients medRxiv https://dx.doi.org/10.1101/2020.03.31.20042333
  5. Paoli, D., Pallotti, F., Colangelo, S., Basilico, F., Mazzuti, L., Turriziani, O., Antonelli, G., Lenzi, A., Lombardo, F. (2020). Study of SARS-CoV-2 in semen and urine samples of a volunteer with positive naso-pharyngeal swab Journal of Endocrinological Investigation https://dx.doi.org/10.1007/s40618-020-01261-1
  6. Feldmann,H, Virus in Semen and the Risk of Sexual Transmission, New Engl J Medicine, 378;15, 2018
  7. Wei, W., Li, Z., Chiew, C., Yong, S., Toh, M., Lee, V. (2020). Presymptomatic Transmission of SARS-CoV-2 — Singapore, January 23–March 16, 2020 Morbidity and Mortality Weekly Report 69(14), 411-415. https://dx.doi.org/10.15585/mmwr.mm6914e1
  8. Baud, D., Greub, G., Favre, G., Gengler, C., Jaton, K., Dubruc, E., Pomar, L. (2020). Second-Trimester miscarriage in a Pregnant Woman With SARS-CoV-2 Infection JAMA 323(21)https://dx.doi.org/10.1001/jama.2020.7233
  9. https://www.sart.org/news-and-publications/news-and-research/press-releases-and-bulletins/asrm-issues-new-guidance-on-fertility-care-during-covid-19-pandemiccalls-for-suspension-of-most-treatments/