Semen collection was performed after 2 to 5 days of sexual abstinence via masturbation in the laboratory.

The semen analysis was performed according to 2021 WHO standards. Concentration and motility were assessed using a Makler counting chamber (Sefi Medical Instruments, Haifa, Israel).

The sperm cytogram was obtained using the RAL kit for the study of sperm morphology, according to the strict Kruger criteria [6].

Sperm DNA fragmentation was determined using the chromatin dispersion test, abbreviated as SCD (Sperm Chromatin Dispersion), also known as the halo test. Fragmentation indices below 15% are considered normal; rates between 15 and 30% are considered moderate; and those above 30% are considered severe. This technique is easy to use, inexpensive, and does not require heavy laboratory equipment. It allows for the analysis of chromatin dispersion under a bright-field optical microscope.

The data were entered using Microsoft Excel 2007. Data analysis was performed using EpiInfo 7.2. Qualitative variables were compared using the chi-square test, and quantitative variables were compared using the Mann-Whitney test. A multivariate binary logistic regression was performed to identify factors associated with high (moderate to severe: >15%) sperm DNA fragmentation. Adjusted odds ratios (OR) were estimated along with their 95% confidence intervals, and statistical significance was assessed using the chi-square test.

We obtained approval from the administrative authorities of the healthcare facility (Authorization No. 03/09/2025), as well as informed consent from the participants, strictly ensuring anonymity.

In our study, the most represented age group was between 40 and 50 years, with a mean age of 47.31 years and a range from 30 to 70 years.

Medical history was characterized by urogenital infections (39%), followed by tobacco use (22%) and alcohol consumption (14.54%). For 53.71% of the participants, the desire to become a father had been present for more than two years.

Abnormalities in the semen analysis primarily involved ejaculate volume and motility, particularly progressive motility, in 81% and 52% of cases, respectively. The sperm cytogram was abnormal in 72% of cases. Morphological abnormalities were predominantly characterized by teratozoospermia in 49% of cases, followed by asthenoteratozoospermia (8%), oligoasthenozoospermia (5%), and asthenozoospermia (4%) (Table 1).

Table 1. Breakdown of patients according to semen analysis/sperm cytogram parameters.

Regarding the sperm DNA fragmentation index, 74 patients had a high fragmentation index that is, moderate to severe fragmentation (>15%) representing 55.31% of the patients. These patients were distributed as follows: 22.73% of patients had severe DNA fragmentation (>30%). Additionally, 32.58% had moderate fragmentation (15% - 30%) (Figure 1).

The Mann-Whitney test was used to compare age with different values of the fragmentation index. Patients older than 50 years had a mean fragmentation index of 33.7%. Thus, age was associated with a high risk of DNA fragmentation, with p = 0.03 (Table 2).

Patients with abnormal semen analysis results had a high (severe or moderate) fragmentation index in 77% of cases. Table 3 shows a comparison between semen analysis parameters, sperm cytogram findings, and sperm DNA fragmentation.

Figure 1. Halosperm, Halotech DNA, SL, Madrid, Spain/SCD: spermatozoa with intact DNA, indicated by green arrows. Orange arrows: spermatozoa with fragmented DNA.

Table 2. Correlation between age and sperm DNA fragmentation.

Table 3. Comparative table of factors associated with high (moderate to severe: >15%) sperm DNA fragmentation.

p < 0.05: significative.

Abnormalities in the semen analysis primarily concerned ejaculate volume and motility, particularly progressive motility, in 81% and 52% of cases, respectively. As for the sperm cytogram, it was abnormal in 72% of cases. Morphological abnormalities were predominantly characterized by teratozoospermia in 49% of cases, followed by asthenoteratozoospermia (8%), oligoasthenozoospermia (5%), and asthenozoospermia (4%). Nevertheless, data from the literature suggest that conventional sperm parameters namely, sperm concentration, motility, and morphology—while serving as the primary tools for assessing male fertility, remain limited and insufficient for predicting sperm functional capacity in the context of diagnosing the causes of male infertility [10].

Furthermore, successful reproduction depends on a key factor: the integrity of sperm DNA. This factor influences not only fertilization but also embryonic development and the ability to carry a pregnancy to term [11].

Regarding the sperm DNA fragmentation index, 74 patients had moderate to severe fragmentation, representing 55.31% of the patients. Patients with abnormal semen analysis results predominantly had an abnormal fragmentation index, although the association was not statistically significant (p = 0.19; Table 2). A study conducted in 2026 found similar results, with a significant link between abnormal semen analysis results and impaired sperm DNA in 1784 patients undergoing assisted reproductive technology (ART) [12]. Furthermore, men with oligozoospermia had a significantly higher DNA fragmentation index than those with normozoospermia, according to a study conducted in 2023 [4]. Additionally, according to the author, embryos derived from sperm with a high fragmentation index did not have a favorable outcome.