Sperm DNA fragmentation (sDF) may be a male factor contributing to recurrent miscarriages and over 50% of infertility cases. ¹ Common semen analysis may miss sperm DNA fragmentation, so if you want to know specifically about your DNA damage, a direct sDF test is needed. However, most doctors disagree on whether testing and treating sDF is helpful because the evidence so far has been mixed. ² A 2019 meta-analysis demonstrated a clear association between sperm DNA damage and recurrent pregnancy losses. ³ In this article, we will cover sperm DNA fragmentation causes, lab tests, and treatments. 

What is sperm DNA fragmentation?

Because sperms need to be compact and agile, most cellular components, including its own DNA repair apparatus, are eliminated. When a sperm leaves its production factory, it naturally sustains some DNA damage. Fortunately, once fertilized, the egg can repair some of the damage before developing into an embryo. ⁴ Sperm DNA fragmentation is a problem when the sperm DNA is so damaged that the egg’s natural DNA repair machinery cannot sufficiently repair it. 

Since the DNA contains genes necessary for healthy fetal development and life sustenance, excessive DNA damage can lead to miscarriage, congenital disorders, and childhood cancers. ⁵⁶ Miscarriages due to sDF tend to occur early, either chemical or within the first 13 weeks. 

Different types of sperm DNA damage

A DNA molecule is present in two strands forming a double helix. When one of the strands breaks, the DNA damage is called a “single strand break” or a nick. Whereas, when both strands break, the DNA damage is called “double strand break.” Other types of DNA damage may include skipping a base pair (basic unit of a DNA polymer) or incorporating an incorrect base pair into the genome. 

The strand breaks are more likely to destabilize the genome, causing infertility and miscarriages. Notably, double-stranded breaks are harder to repair and more likely to disrupt embryonic development. 

Typically, our DNA wraps around a protein called histones to form chromosomes. But to further compact the DNA into the sperm head, histones are exchanged for another protein called protamine, which can better compact the sperm genomes into donut-shaped structures. This process is called “protamination.” These donuts have regions that are linked to packing strings (nuclear matrix) inside the sperm head. ⁸ 

What causes sperm DNA fragmentation?

DNA damage can occur both during sperm production and transport. However, most of the DNA damage that contributes to sDF happens as the semen travels out of the testes into the semen tube (epididymis) prior to ejaculation. 

Single strand breaks are caused by reactive oxygen species, such as environmental toxicants, smoking, alcohol, diet, radiation, infection, varicocele, or even the sperm’s own metabolism. ¹¹¹² Double-strand breaks are more likely caused by attempted cellular suicide (apoptosis) during the sperm production process or by enzymes triggered by heat and oxidative stress. 

Factors that influence sperm DNA damage and are indications for testing sperm DNA fragmentation include: ¹³¹⁴¹⁵ 

• Infection
• High fever and elevated testicular temperature
• Varicocele (enlargement of the veins in the scrotal sac)
• Advanced paternal age (>30)
• Obesity and Poor diet
• Drug use and Cigarette smoking
• Exposure to environmental and occupational pollutants
• Increased length of sexual abstinence
• Stress
• Alcohol abuse

Sperm and semen factors that may increase sDF include:

• Reduced antioxidants in semen and sperm cells
• Poor sperm DNA packaging, including due to protamine deficiency ¹⁶

What are sperm DNA fragmentation symptoms?

Most men are unaware that they have sDF until they try to have children because there are no symptoms. sDF contributes to male infertility, failed assisted reproduction technique (ART, including IVF) treatment cycles, or recurrent miscarriage. ¹⁷ 

High DNA fragmentation is associated with reduced sperm count, motility, and morphology. ¹⁸¹⁹ But it is totally possible to have sDF with normal semen analysis results, so you may need specific tests for sDF if you struggle with infertility and recurrent pregnancy loss. ²⁰ Keep in mind, also, that they don’t routinely test for sDF as doctors disagree on its usefulness, so you may want to ask your doctor about it.

sDF destabilizes the embryonic genome, which can stop development and disturb implantation, leading to early miscarriages. ²¹ The paternal genome is switched on from day two of development and that is when sDF is more likely to affect embryonic development. ²² DNA damage in the embryo could also lead to cell degeneration and genetic mutations, causing congenital genetic diseases and childhood cancers. ²³²⁴²⁵

sDF is associated with the following negative pregnancy outcomes: ²⁶ 

• Miscarriage ²⁷²⁸
• Recurrent pregnancy loss ²⁹³⁰
• Reduced live birth rates ³¹³²

Sperm DNA fragmentation diagnosis

Semen analysis

Semen analysis (also called spermogram or seminogram) evaluates sperm appearance and semen composition, such as semen volume, pH, sperm concentration, motility, vitality, and morphology. ³³ Subfertile men with abnormal sperm parameters do have more sDF than fertile men. ³⁴ However, while defective genetic material is associated with abnormal sperm, this test does not directly test for genetic defects. 

Almost 15% of infertile men have normal semenalysis, according to the WHO 2010. ³⁵ Interestingly, these patients with normal sperm parameters can also have high sperm DNA fragmentation. 

Specific sDF tests

sDF tests are controversial because some studies showed that sDF correlates with positive ICSI outcomes, but others show the opposite result. ³⁶³⁷ The conflicting results and lack of large clinical studies led the American Society for Reproductive Medicine to recommend against these tests. ³⁸ In addition, there isn’t an established threshold for sDF test values or a standardized treatment protocol. ³⁹ Therefore, many fertility specialists only check for sDF once they already rule out all the other causes, but some prefer to test it early on.

Indications for sDF tests include: ⁴⁰ 

• Unexplained infertility
• Poor embryo development
• Multiple failed IVF/ICSI treatment
• Recurrent miscarriage 
• Advanced age (>30 years)
• Varicocele 
• Poor semen parameters (spermogram)
• Exposure to harmful substances

The most common tests include the Comet assay, the terminal transferase dUTP nick end labeling (TUNEL) assay, Sperm Chromatin Structure Assay (SCSA), and the Sperm Chromatin Dispersion (SCD or Halo) test. ⁴¹ These tests are not created equal because each measures different types of DNA damage (See table). ⁴²⁴³

Test	What it measures	Types of DNA damage detected	Caveats
TUNEL	The number of nicked and free DNA ends	Single-strand breaks Extensive double-strand breaks	Does not detect double-strand breaks in regions that attach to nuclear matrix
SCSA	Broken DNA is more likely to denature and take up red dye. Unbroken DNA emits green color.	Single strand breaks Extensive double strand breaks	Does not detect double-strand breaks in regions that attach to nuclear matrix. Is an indirect test. May measure more susceptibility to DNA damage than actual damage 44
SCD	Fragment size of DNA denatured into single-strands with chemical or heat	Single-strand breaks Extensive double strand breaks	Does not detect double-strand breaks in regions that attach to nuclear matrix
Neutral Comet	Amount of double-stranded fragmented DNA	All double strand breaks, including in regions that attach to nuclear matrix
Alkaline Comet	Amount of double and single strand fragmented DNA	All double and single strand breaks
Two-tailed Comet	Combination of neutral and alkaline Comet	Single-strand breaks Extensive double strand breaks	Less standardized test
Toluidine blue staining	Overall DNA integrity inside sperm heads		Fast but non-DFI test
CMA3 staning	Sperm DNA protamination		Non-DFI test, value >30% lowers ICSI fertilization rate 45

TUNEL, SCSA, and SCD results typically correlate with alkaline Comet but not neutral Comet results. This is because only the neutral Comet can detect localized strand breaks between the donut structures that attach to the nuclear matrix. ⁴⁶ 

Reference ranges

These tests measure DNA damage and display results as DNA Fragmentation Index (DFI), which shows the percentage of cells with seriously damaged DNA. ⁴⁷ 

For instance, reference levels in SCSA are following: ⁴⁸  

• ≤ 15% DFI = excellent to good sperm DNA integrity 
• > 15 to < 25% DFI = good to fair sperm DNA integrity
• > 25 to < 50% DFI = fair to poor sperm DNA integrity
• ≥ 50% DFI = very poor sperm DNA integrity

With the TUNEL technique, studies have shown that DFI >20% indicates poor sperm DNA quality, and can be used to distinguish between fertile and infertile men. ⁴⁹ 

Treatments for Sperm DNA fragmentation

Lifestyle Changes

sDF has a major lifestyle contributor, so treating sDF involves diet and lifestyle changes to reduce oxidative stress and DNA damage in sperm cells, including: ⁵⁰⁵¹ 

• Fever reduction with antipyretics (paracetamol, ibuprofen)
• Treating the infections
• Quit drugs, smoking and occupational hazards (vibrations, Heavy metals, PVC) 
• Diet – fresh foods, particularly those containing antioxidants and vitamin C & E
• Avoiding toxic exposures
• Varicocele surgery
• Frequent ejaculation 

Lifestyle risk factors	Lifestyle modifications
Smoking (Nicotine, Cd, Pb, Benzo(a)-pyrene)- Oxidative stress causes significant DNA fragmentations and spermatozoids are especially susceptible to it 52	Cessation of smoking
Air pollution (Polycyclic aromatic hydrocarbons and air particulate matter) can damage DNA 53	Avoid spending time outside in high pollution hours, wear face masks, and spend more time in nature.
POP: organochlorine pesticides, such as DDT, industrial chemicals, polychlorinated biphenyls (PCB), unintentional by-products of many industrial processes, such as polychlorinated dibenzo-p-dioxins (PCDD) and dibenzofurans (PCDF) or ‘dioxins’ 54	PCB accumulates in the ends of the food chain: Avoid fatty fish, especially farmed.
Infection – Overall inflammation causes oxidative stress leading to damage DNA in spermatozoa 55	Antibiotics and antipyretics (this includes something so seemingly irrelevant such as bad teeth)
Testicular heat – Increase in DNA fragmentation with 2–3°C temperature increase 56	Avoid cycling with tight pants, sauna use, using laptops on closed legs
Mobile phone radiation – increased levels of SDF and decreased antioxidants [39,40] 5758	Do not store mobile phone in trouser pocket and/or consider radiation-blocking underwear
UV radiation – Sperm genetic material is at higher risk of DNA fragmentation due to UV radiation (254 nm) 59	Avoid professional hazards
Caffeine – Some authors believe it might cause higher SDF, especially in unprotaminated regions, but there is no consensus 60	Reduce coffee intake
Alcohol – Alcohol may increase the percentage of spermatozoa with DNA fragmentation and apoptosis 61	Reduce alcohol intake to one unit maximum per day, preferably as red wine, because it contains antioxidants
Drugs – Ecstasy and opiates create oxidative stress metabolites 62	Cessation of drug intake
Heavy metals – Increase in percentage of spermatozoa with DNA fragmentation 63	Avoid occupational exposure and smoking or exposure to cigarette smoke. Avoid fish with high mercury levels.
Bisphenol A and PVC – Linked with semen quality, DNA integrity, oxidative stress in infertile men 64	Avoid plastic packaging, canned foods. Don’t heat or store food in plastic containers.
Obesity – Poor spermatogenesis and associated with high SDF 65	Losing weight through healthy diet and exercise
Imbalanced diet – Lack of antioxidants needed to repair DNA damage 66	Better diet. Include more foods rich in vitamins E and C such as fruits and vegetables, e.g. pepper, cabbage, fish, and nuts.  Oxidant supplements such as vitamins E and C, selenium, coenzyme Q10, N-acetylcysteine, zinc, and L-carnitine
Abstinence – Supposedly leads to accumulation of spermatozoids with DNA that hasn’t been repairing DNA 67	Frequent ejaculation, more than once a day 68

After three months of these initiatives to reduce sDF, it is a good idea to retest for sDF levels to see if the changes are working. ⁶⁹

Fertility Treatments

sDF testing may have very important predictive value for fertility treatments. Couples with low sperm DNA damage can succeed with traditional intrauterine insemination (IUI) or in vitro fertilization (IVF). Whereas, when DFI values are higher, directly injecting sperms into the cytoplasm of eggs (Intracytoplasmic Sperm Injection, ICSI) instead of traditional IVF may increase the chance of success. ⁷⁰⁷¹ Traditional IVF exposes the egg to the sperms in the test tube for 90 minutes, whereas ICSI directly injects the sperm into the oocyte. Therefore, ICSI might reduce the amount of time both the eggs and sperms are exposed to reactive oxygen species. ⁷² ICSI typically costs more than IVF, and may also confer some health risks in the offspring, ⁷³ so ICSI is usually considered only after IVF is unsuccessful. 

Typically, natural ejaculates are noninvasively collected and used for IVF and ICSI. In men with sDF, most of the sperm DNA damage that contribute to infertility and miscarriage happens once the semen leaves the testicle for an ejaculation. ⁷⁴ Therefore, direct aspiration of sperm from the testicle (testicular sperm aspiration, TESA) may improve the chance of a successful pregnancy than ejaculate for these men. ⁷⁵ However, TESA is not without risk—it is an invasive surgical procedure with small chances of complications, including infection, bleeding, and testicular atrophy. ⁷⁶ In addition, TESA combined with ICSI may increase the chance of chromosomal abnormalities in the embryo and offspring. ⁷⁷ 

Can you have a successful pregnancy with sDF?

Fortunately, couples where the male partner has a high percentage of sperm DNA fragmentation have had successful pregnancies. Your chances will be higher if the percentage of sperm bearing high levels of SDF is lower. The chances of successful pregnancies are also greater if the spermatozoids with high DNA fragmentation fertilize younger oocytes, which are much more efficient at repairing sperm DNA damages. ⁷⁸