Abstract
Infertility affects approximately 15% of couples of reproductive age worldwide, with male factors contributing to nearly 40%–50% of cases. Because semen quality is a fundamental indicator of male fertility, understanding its determinants is essential for clinical assessment. In this retrospective cohort study involving 131 183 patients, we compared semen parameters between infertile men and healthy controls and explored the relationships among age, infectious microorganisms, and semen quality. Our analysis demonstrated that infertile men had significantly poorer semen quality than controls did. Age also played a critical role, with the lowest risk of male infertility observed in the 25–29 years age group; across all age groups, risk followed a U-shaped pattern. Furthermore, the presence of antisperm antibodies, Neisseria gonorrhea, Chlamydia trachomatis, Ureaplasma urealyticum, and/or Mycoplasma genitalium was associated with reduced semen quality and increased infertility risk. In contrast, although herpes simplex virus II was detected in some semen samples, its overall prevalence was low, and no significant impact on semen parameters was detected. These findings emphasize the complex interplay among age, infection, and immune factors in shaping male reproductive potential. They also highlight the need for targeted prevention and treatment strategies to mitigate the burden of male infertility and to improve future therapeutic outcomes.
Keywords: antisperm antibodies, Chlamydia trachomatis, male fertility, Mycoplasma genitalium, Neisseria gonorrhea, semen, Ureaplasma urealyticum
INTRODUCTION
Infertility is a significant health concern for couples of reproductive age, affecting approximately 15% of couples worldwide, with male factors contributing to 40%–50% of these cases.1 Consequently, male infertility not only has direct health implications for men but also substantially affects the physiological and psychological well-being of both partners.2 As such, infertility has emerged as a major public health challenge with broad societal implications.3
Semen quality is the primary indicator for assessing male fertility.4,5 Key semen parameters, including liquefaction time, pH, sperm concentration, motility, and the proportion of morphologically abnormal spermatozoa, serve as critical markers in the evaluation of male infertility.5 Abnormal sperm morphology not only reduces fertility potential but also, in severe cases, can result in teratozoospermia, a major cause of male infertility.6 Clinically, semen analysis results are widely utilized as an essential diagnostic tool in the assessment of male reproductive function.
With delayed childbearing and the implementation of China’s three-child policy, the proportion of older couples attempting conception has gradually increased.7 Aging in men is associated with decreases in semen volume, total sperm count, and overall motility, which are associated with lower clinical pregnancy and live birth rates.4,5 Male reproductive tract infections are another critical contributor to infertility and have garnered increasing clinical attention.8 Pathogens such as Chlamydia trachomatis (CT), Neisseria gonorrhea (NG), Mycoplasma genitalium (MG), and Ureaplasma urealyticum (UU) are commonly transmitted via sexual contact and can cause urethritis, epididymitis, prostatitis, and, in females, vaginitis, endometritis, and pelvic inflammatory disease.9,10 These infections are also linked to infertility, subfertility, and adverse pregnancy outcomes.11,12 Subclinical or untreated infections may ascend along the reproductive tract, persist asymptomatically, and increase the risk of transmission to sexual partners, posing significant reproductive health risks.13,14 However, the specific effects of these pathogens on semen quality and their associations with male infertility remain underexplored.14 Immune-mediated factors account for 15%–30% of male infertility cases.15 Normally, the blood–testis and blood–epididymis barriers prevent the production of antisperm antibodies (AsAb).16 Nonetheless, conditions such as epididymitis, testicular trauma, vasal obstruction, or testicular inflammation can expose sperm antigens to the immune system, triggering AsAb production.17 Binding of AsAb to spermatozoa can impair sperm function and compromise male fertility.17
Therefore, the present study comprehensively evaluated semen quality in infertile men and investigated the effects of UU, CT, NG, MG, and AsAb on semen parameters, as well as their associations with the risk of male infertility.
PARTICIPANTS AND METHODS
Participants
This retrospective cohort study involved male patients who sought medical care at the Chongqing Health Center for Women and Children (Chongqing, China) between January 2017 and November 2024. The study obtained approval from the Ethics Committee of the Chongqing Health Center for Women and Children (Approval No. 2024 Ethics Department 052), and informed consent was obtained from all participants prior to data collection. All methods were carried out in accordance with the relevant ethical guidelines and regulations.18 The datasets used and/or analyzed during the current study are available from the corresponding author upon reasonable request.
Inclusion and exclusion criteria
The inclusion criteria were as follows: (1) availability of complete clinical data; (2) were males of reproductive age; (3) had a history of regular sexual activity; (4) did not use antimicrobial drugs during the 2-week preceding semen testing; and (5) had no history of successful pregnancies with their partners. The exclusion criteria included the following: (1) recent fever, sleep deprivation, or similar conditions; (2) recent severe acute illnesses; or (3) secondary male infertility caused by conditions such as varicocele, vas deferens obstruction, or erectile dysfunction.
Subgroups
The patients were divided into two groups on the basis of their diagnoses: the control group and the infertility group. The inclusion criterion for the control group was that the couple had previously conceived naturally and successfully delivered a child. For the infertility group, the inclusion criteria were male patients who had engaged in regular sexual activity for more than 1 year without using contraceptive measures but were unable to achieve natural conception due to male-related factors.
Semen collection and routine detection
Before semen collection, patients were instructed to abstain from ejaculation for 2–7 days. All semen samples were obtained by masturbation in a private room adjacent to the andrology laboratory within the Chongqing Health Center for Women and Children. Immediately after ejaculation, the samples were kept at body temperature (approximately 37°C) either by holding the specimen container in the hand or by placing it in a prewarmed incubator. The samples were delivered to the laboratory within 30 min after production. The analysis followed the 6th edition of the World Health Organization (WHO) manual.18 The semen volume was measured by weighing the sample after complete liquefaction (30–60 min). A 10-μl sample of liquefied semen was placed on a Makler semen counting chamber (Sefi Medical Instruments, Haifa, Israel), and the total number of spermatozoa (TS) in 10 consecutive squares was counted under a phase-contrast microscope (CX41; Olympus, Tokyo, Japan) at 200× magnification. The total sperm count was determined as TS × semen volume × 106, and the sperm concentration was calculated as TS × 106 ml−1. The hypo-osmotic swelling test was used to assess sperm viability, while modified Papanicolaou staining was used for manual evaluation of the morphology of at least 200 spermatozoa, and the percentage of normal sperm was calculated. The teratozoospermia index (TZI) was defined as the total number of identified morphological defects divided by the number of spermatozoa presenting at least one defect. The sperm deformity index (SDI) was calculated as the total number of identified morphological defects divided by the total number of spermatozoa assessed.
Detection of AsAb, NG, CT, UU, MG, and herpes simplex virus II (HSV-II)
AsAb was detected using an AsAb kit, which is based on the mixed antiglobulin agglutination (MAR) method, with reagents supplied by Anhui Anke Biotechnology Group Co., Ltd. (Hefei, China). CT, UU, MG, NG, and HSV-II were detected using a nucleic acid detection kit based on polymerase chain reaction (PCR)-fluorescent probes, with reagents provided by Hunan Shengxiang Biotechnology Co., Ltd. (Changsha, China). The testing procedures strictly adhered to the protocols outlined in the respective reagent kits.
Statistical analyses
Data were analyzed with Stata 17 (StataCorp LLC, College Station, TX, USA). To avoid narrow confidence intervals due to the large sample size, we converted all continuous variables into binary variables, and a Chi-square test was used. The reference ranges for semen parameter values from the 6th edition of the WHO manual18 were used. The semen parameters were then classified as positive or negative based on the reference range provided (Supplementary Table 1).
Supplementary Table 1.
The confidence interval for semen parameters
| Semen parameters | Reference range |
|---|---|
| Semen volume | ≥1.4 ml |
| PH value | ≥7.2 |
| Sperm concentration | ≥16×106/ml |
| Sperm vitality | ≥54% |
| Progressive sperm motility | ≥30% |
| Total sperm count | ≥39×106/ml |
| Total motility | ≥42% |
| Normal sperm (stained) | ≥4% |
| Teratozoospermia index | 1.0–1.6 |
| Sperm deformity index | 1.0–1.6 |
In each analysis, the semen parameters were classified into two groups on the basis of the patients’ diagnosis: the infertility group and the control group. Chi-square tests were used for univariate analysis, and the infertility risk associated with each parameter was calculated separately. Logistic regression and Poisson regression were used for multivariate regression analysis, adjusting for potential confounders, including diagnosis, age, abstinence time, semen volume, year, and pH. The adjusted odds ratio (OR) and 95% confidence intervals (CIs) were obtained from the logistic regression model, whereas the adjusted relative risk ratio (RR) and its 95% CI were obtained from the Poisson regression model. These models were used to determine the occurrence risk of male infertility and the prevalence risk of genital pathogens. Statistical significance was confirmed only when both models yielded consistent results.
To assess the prevalence of microbial infections and their association with semen quality, Chi-square tests were used to compare the positivity rates of NG, CT, UU, MG, and HSV-II from 2017 to 2024. To ensure the robustness of the findings, we further performed multivariable logistic regression and multivariable Poisson regression, adjusting for potential confounders such as age and year of testing, to evaluate the infection rates of these genital pathogens among infertile men. P < 0.05 was considered to indicate a statistically significant difference.
RESULTS
Patient characteristics
Between January 2017 and November 2024, a total of 376 205 patients underwent semen testing at the Chongqing Health Center for Women and Children. After excluding duplicate tests, 131 183 patients were included in the statistical analysis. Patients were stratified into two groups according to clinical diagnosis: the control group (n = 62 053, age [mean±standard deviation, s.d.]: 32.1 ± 5.3 years) and the infertility group (n = 69 130, age [mean ± s.d.]: 32.9 ± 5.8 years), as shown in Supplementary Figure 1 (103.4KB, tif) . In the overall study population, the incidence of male infertility was 52.7%. Patients were further categorized into six age groups: <25 years, 25–29 years, 30–34 years, 35–39 years, 40–44 years, and ≥45 years. The greatest proportions of semen analyses were recorded in the age groups of 25–29 years and 30–34 years, accounting for 30.2% and 38.0% of the cases, respectively (Supplementary Figure 2a (57.9KB, tif) ). Infertility risk was the lowest in the 25–29 years age group, which was used as the reference category. Risk analysis revealed a U-shaped pattern, with infertility risk progressively increasing after age of 29 years (Supplementary Figure 2b (57.9KB, tif) and Supplementary Table 2). When stratified by testing year (2017–2024), the infertility rate peaked in 2020 (59.7%) and declined annually thereafter (Figure 1a and Supplementary Table 2).
Table 1.
The male infertility patients classified by semen parameter values (n=131 183)
| Variable | Univariate analysis | Multivariable logistic regression | Multivariable Poisson regression | |||||
|---|---|---|---|---|---|---|---|---|
|
|
|
|
||||||
| Infertility | Total | Infertility rate (%) | a P | Adjusted OR (95% CI) | b P | Adjusted RR (95% CI) | c P | |
| Overall | 69 130 | 131 183 | 52.7 | - | - | - | - | - |
| Semen volume | ||||||||
| N (≥1.4 ml) | 65 400 | 125 129 | 52.3 | Reference | Reference | Reference | Reference | Reference |
| A (<1.4 ml) | 3730 | 6054 | 61.6 | <0.001 | 1.47 (1.39–1.55) | <0.001d | 1.17 (1.13–1.21) | <0.001d |
| pH | ||||||||
| N (≥7.2) | 61 773 | 115 673 | 53.4 | Reference | Reference | Reference | Reference | Reference |
| A (<7.2) | 7357 | 15 510 | 47.4 | <0.001 | 0.75 (0.73–0.78) | <0.001e | 0.88 (0.86–0.90) | <0.001e |
| Total sperm count | ||||||||
| N (≥39×106 per ejaculate) | 63 889 | 122 903 | 52.0 | Reference | Reference | Reference | Reference | Reference |
| A (<39×106 per ejaculate) | 5241 | 8280 | 63.3 | <0.001 | 1.54 (1.47–1.62) | <0.001f | 1.19 (1.15–1.22) | <0.001f |
| Total motility | ||||||||
| N (≥42%) | 57 780 | 112 953 | 51.2 | Reference | Reference | Reference | Reference | Reference |
| A (<42%) | 11 350 | 18 230 | 62.3 | <0.001 | 1.56 (1.50–1.61) | <0.001f | 1.19 (1.17–1.21) | <0.001f |
| Sperm vitality | ||||||||
| N (≥54%) | 62 280 | 123 880 | 50.3 | Reference | Reference | Reference | Reference | Reference |
| A (<54%) | 4850 | 7303 | 66.4 | <0.001 | 1.85 (1.76–1.95) | <0.001f | 1.27 (1.23–1.30) | <0.001f |
| Sperm concentration | ||||||||
| N (≥16×106 ml−1) | 58 215 | 115 787 | 50.3 | Reference | Reference | Reference | Reference | Reference |
| A (<16×106 ml−1) | 10 915 | 15 396 | 70.9 | <0.001 | 2.38 (2.29–2.47) | <0.001f | 1.37 (1.34–1.40) | <0.001f |
| Normal spermatozoa (stained) | ||||||||
| N (≥4%) | 28 875 | 54 777 | 52.7 | Reference | Reference | Reference | Reference | Reference |
| A (<4%) | 40 255 | 76 406 | 52.7 | 0.920 | 1.09 (1.06–1.11) | <0.001f | 1.04 (1.02–1.06) | <0.001f |
| Progressive sperm motility | ||||||||
| N (≥30%) | 63 496 | 122 166 | 52.0 | Reference | Reference | Reference | Reference | Reference |
| A (<30%) | 5634 | 9017 | 62.5 | <0.001 | 1.70 (1.63–1.79) | <0.001f | 1.23 (1.20–1.27) | <0.001f |
| Teratozoospermia index | ||||||||
| N (≤1.6) | 37 717 | 72 784 | 51.8 | Reference | Reference | Reference | Reference | Reference |
| A (>1.6) | 31 413 | 58 399 | 53.8 | <0.001 | 1.01 (0.98–1.03) | 0.565f | 1.00 (0.99–1.02) | 0.720f |
| Sperm deformity index | ||||||||
| N (≤1.6) | 38 549 | 74 036 | 52.1 | Reference | Reference | Reference | Reference | Reference |
| A (>1.6) | 30 581 | 57 147 | 53.5 | <0.001 | 0.99 (0.96–1.01) | 0.296f | 0.99 (0.98–1.01) | 0.450f |
aP value of univariate analysis. bP value of multivariable logistic regression. cP value of multivariable Poisson regression. dAdjusted for diagnosis, age, abstinence time, year, and pH value. eAdjusted for diagnosis, age, abstinence time, semen volume, and year. fAdjusted for diagnosis, age, abstinence time, semen volume, year, and pH value. RR: risk ratio; OR: odds ratio; -: not calculable in the overall population; N: normal result; A: abnormal result; CI: confidence interval
Figure 1.
Annual incidence and clearance of urogenital pathogens in infertile men. (a) Annual incidence of infertile men with semen tests. (b) Impact of the clearance of NG, CT, UU, and MG on male infertility. *P < 0.05. AsAb: antisperm antibodies; NG: Neisseria gonorrhea; CT: Chlamydia trachomatis; UU: Ureaplasma urealyticum; MG: Mycoplasma genitalium; HSV-II: herpes simplex virus II; TN: turned negative; not TN: not turned negative.
Differences in semen parameter values between the infertility and control groups
Overall, more than half of the infertile men had abnormal semen parameter values. Compared with controls, infertile men had significantly greater abnormal semen volume (61.6%), total sperm count (63.3%), total motility (62.3%), sperm vitality (66.4%), sperm concentration (70.9%), normal sperm (stained: 52.7%), and progressive motility (62.5%). These differences remained statistically significant after multivariable adjustment. Notably, sperm concentration was the highest among all parameters. Both logistic regression (OR = 2.38; 95% CI: 2.29–2.47; P < 0.001) and Poisson regression (RR = 1.37; 95% CI: 1.34–1.40; P < 0.001) consistently revealed significantly higher abnormality rates in infertile men than those in controls. In contrast, no significant differences were found in the teratozoospermia index or sperm deformity index in the multivariable analyses (both P > 0.05; Table 1).
Prevalence of genital pathogens and AsAb
We assess the prevalence of microbial infections and their association with semen quality. The infection rates of CT (3.3% vs 1.2%; logistic regression: OR = 6.92, 95% CI: 6.31–7.59, P < 0.001; Poisson regression: RR = 2.65, 95% CI: 2.51–2.79, P < 0.001) and UU (22.6% vs 18.6%; logistic regression: OR = 1.57, 95% CI: 1.36–1.80, P < 0.001; Poisson regression: RR = 1.42, 95% CI: 1.26–1.60, P < 0.001) were significantly greater in the infertility group than those in the control group (Figure 2). The prevalence of AsAb was also significantly greater in infertile men than that in controls (2.9% vs 2.3%, P < 0.001). Logistic regression indicated a modest but statistically significant association with infertility (P = 0.039), whereas Poisson regression did not confirm this finding (P = 0.199). Thus, the role of AsAb in male infertility warrants further investigation (Figure 2).
Figure 2.
Prevalence of Neisseria gonorrhea, Chlamydia trachomatis, Ureaplasma urealyticum, Mycoplasma genitalium, antisperm antibody, and Herpes simplex virus II in the overall population. aP value of univariate analysis. bP value of multivariable logistic regression. cP value of multivariable Poisson regression. dAdjusted for diagnosis, age, abstinence time, semen volume, year, and pH value. CG: control group; IG: infertility group; RR: risk ratio; OR: odds ratio; -: not calculable.
Effects of genital pathogens and AsAb on semen quality
We then examined the effects of infections and AsAb on semen quality. CT infection significantly increased the abnormal rates of sperm vitality, normal sperm (stained), the teratozoospermia index, and the sperm deformity index (all P < 0.05; Table 2). UU infection was initially associated with abnormalities in total motility, normal sperm (stained), the teratozoospermia index, and the sperm deformity index, but these associations were not significant after multivariable adjustment (all P > 0.05; Supplementary Table 3). NG infection was strongly associated with abnormalities in total sperm count, total motility, sperm vitality, sperm concentration, and progressive motility (all P < 0.001; Supplementary Table 4). MG infection significantly increased the rates of abnormal pH value, total sperm count, and sperm concentration (all P < 0.05; Supplementary Table 5). AsAb were significantly associated with higher abnormal rates of total motility, sperm vitality, progressive motility, pH value, the teratozoospermia index, and the sperm deformity index (all P < 0.001; Supplementary Table 6).
Table 2.
The effect of Chlamydia trachomatis on semen quality in the overall population
| Variable | Abnormal (A) | Normal (N) | Total | Abnormal rate (%) | a P | Multivariable logistic regression | Multivariable Poisson regression | ||
|---|---|---|---|---|---|---|---|---|---|
|
|
|
||||||||
| Adjusted OR (95% CI) | b P | Adjusted RR (95% CI) | c P | ||||||
| Semen volume (N ≥1.4 ml; A <1.4 ml) | |||||||||
| Total | 2848 | 67 463 | 70 311 | 4.1 | - | - | - | - | - |
| CT− | 2722 | 65 245 | 67 967 | 4.0 | Reference | Reference | Reference | Reference | Reference |
| CT+ | 126 | 2218 | 2344 | 5.4 | 0.001 | 1.21 (1.00–1.46) | 0.050d | 1.19 (0.99–1.43) | 0.057d |
| pH (N ≥7.2; A <7.2) | |||||||||
| Total | 6924 | 63 387 | 70 311 | 9.8 | - | - | - | - | - |
| CT− | 6704 | 61 263 | 67 967 | 9.9 | Reference | Reference | Reference | Reference | Reference |
| CT+ | 220 | 2124 | 2344 | 9.4 | 0.445 | 1.12 (0.97–1.30) | 0.127d | 1.11 (0.97–1.28) | 0.143d |
| Total sperm count (N ≥39×106 per ejaculate; A <39×106 per ejaculate) | |||||||||
| Total | 3471 | 66 840 | 70 311 | 4.9 | - | - | - | - | - |
| CT− | 3328 | 64 639 | 67 967 | 4.9 | Reference | Reference | Reference | Reference | Reference |
| CT+ | 143 | 2201 | 2344 | 6.1 | 0.008 | 1.05 (0.88–1.26) | 0.566d | 1.05 (0.89–1.24) | 0.577d |
| Total motility (N ≥42%; A <42%) | |||||||||
| Total | 8223 | 62 088 | 70 311 | 11.7 | - | - | - | - | - |
| CT− | 7891 | 60 076 | 67 967 | 11.6 | Reference | Reference | Reference | Reference | Reference |
| CT+ | 332 | 2012 | 2344 | 14.2 | <0.001 | 1.00 (0.88–1.13) | 0.993d | 1.00 (0.89–1.13) | 0.947d |
| Sperm vitality (N ≥54%; A <54%) | |||||||||
| Total | 3116 | 67 195 | 70 311 | 4.4 | - | - | - | - | - |
| CT− | 2946 | 65 021 | 67 967 | 4.3 | Reference | Reference | Reference | Reference | Reference |
| CT+ | 170 | 2174 | 2344 | 7.3 | <0.001 | 1.24 (1.05–1.46) | 0.013d | 1.21 (1.03–1.41) | 0.019d |
| Sperm concentration (N ≥16×106 ml−1; A <16×106 ml−1) | |||||||||
| Total | 6084 | 64 227 | 70 311 | 8.7 | - | - | - | - | - |
| CT− | 5792 | 62 175 | 67 967 | 8.5 | Reference | Reference | Reference | Reference | Reference |
| CT+ | 292 | 2052 | 2344 | 12.5 | <0.001 | 1.13 (0.99–1.29) | 0.064d | 1.12 (0.99–1.27) | 0.076d |
| Normal spermatozoa (stained; N ≥4%; A <4%) | |||||||||
| Total | 27 296 | 43 015 | 70 311 | 38.8 | - | - | - | - | - |
| CT− | 26 051 | 41 916 | 67 967 | 38.3 | Reference | Reference | Reference | Reference | Reference |
| CT+ | 1245 | 1099 | 2344 | 53.1 | <0.001 | 1.58 (1.28–1.95) | <0.001d | 1.55 (1.27–1.90) | <0.001d |
| Progressive sperm motility (N ≥30%; A <30%) | |||||||||
| Total | 4097 | 66 214 | 70 311 | 5.8 | - | - | - | - | - |
| CT− | 3883 | 64 084 | 67 967 | 5.7 | Reference | Reference | Reference | Reference | Reference |
| CT+ | 214 | 2130 | 2344 | 9.1 | <0.001 | 1.14 (0.98–1.33) | 0.084d | 1.13 (0.98–1.30) | 0.097d |
| Teratozoospermia index (N ≤1.6; A >1.6) | |||||||||
| Total | 29 004 | 41 307 | 70 311 | 41.3 | - | - | - | - | - |
| CT− | 27 860 | 40 107 | 67 967 | 41.0 | Reference | Reference | Reference | Reference | Reference |
| CT+ | 1144 | 1200 | 2344 | 48.8 | <0.001 | 1.25 (1.13–1.38) | <0.001d | 1.22 (1.11–1.34) | <0.001d |
| Sperm deformity index (N ≤1.6; A >1.6) | |||||||||
| Total | 28 616 | 41 695 | 70 311 | 40.7 | - | - | - | - | - |
| CT− | 27 482 | 40 485 | 67 967 | 40.4 | Reference | Reference | Reference | Reference | Reference |
| CT+ | 1134 | 1210 | 2344 | 48.4 | <0.001 | 1.26 (1.14–1.39) | <0.001d | 1.22 (1.11–1.35) | <0.001d |
aP value of univariate analysis. bP value of multivariable logistic regression. cP value of multivariable Poisson regression. dAdjusted for diagnosis, age, abstinence time, and year. CT: Chlamydia trachomatis; +: positive; −: negative; RR: risk ratio; OR: odds ratio; -: not calculable
Supplementary Table 2.
Basic characteristics of the overall sample (n=131 183)
| Variables | Univariate analysis | Multivariate analysis | ||||||
|---|---|---|---|---|---|---|---|---|
|
|
|
|||||||
| Infertility | Total | Infertility rate (%) | P a | Logistic regression | Poisson regression | |||
|
|
|
|||||||
| Adjusted OR (95% CI) | P b | Adjusted RR (95% CI) | P c | |||||
| Overall | 69 130 | 131 183 | 52.7 | - | - | - | - | - |
| Age (years) | ||||||||
| <25 | 2064 | 3560 | 58.0 | <0.001 | <25 (<0.001) | <0.001d | 1.15 (1.10–1.21) | <0.001d |
| 25–29 | 18 967 | 39 603 | 479 | Reference | Reference | Reference | Reference | Reference |
| 30–34 | 25 867 | 49 906 | 51.8 | <0.001 | 1.18 (1.15–1.21) | <0.001d | 1.08 (1.06–1.10) | <0.001d |
| 35–39 | 13 159 | 23 118 | 56.9 | <0.001 | 1.42 (1.37–1.47) | <0.001d | 1.17 (1.15–1.20) | <0.001d |
| 40–44 | 5861 | 9837 | 59.6 | <0.001 | 1.46 (1.39–1.53) | <0.001d | 1.18 (1.15–1.21) | <0.001d |
| ≥45 | 3212 | 5159 | 62.3 | <0.001 | 1.57 (1.48–1.67) | <0.001d | 1.21 (1.17–1.26) | <0.001d |
| Year | ||||||||
| 2017 | 10 952 | 19 245 | 56.9 | Reference | Reference | Reference | Reference | Reference |
| 2018 | 11 603 | 19 398 | 59.8 | <0.001 | 1.13 (1.08–1.17) | <0.001e | 1.05 (1.02–1.08) | <0.001e |
| 2019 | 10 512 | 17 824 | 59.0 | <0.001 | 1.14 (1.09–1.19) | <0.001e | 1.06 (1.03–1.08) | <0.001e |
| 2020 | 8947 | 14 993 | 59.7 | <0.001 | 1.17 (1.12–1.22) | <0.001e | 1.07 (1.04–1.10) | <0.001e |
| 2021 | 8612 | 16 059 | 53.6 | <0.001 | 0.89 (0.86–0.93) | <0.001e | 0.95 (0.93–0.98) | <0.001e |
| 2022 | 7169 | 14 360 | 49.9 | <0.001 | 0.76 (0.73–0.80) | <0.001e | 0.88 (0.86–0.91) | <0.001e |
| 2023 | 8003 | 17 149 | 46.7 | <0.001 | 0.66 (0.63–0.69) | <0.001e | 0.82 (0.80–0.85) | <0.001e |
| 2024 | 3332 | 12 155 | 27.4 | <0.001 | 0.23 (0.22–0.24) | <0.001e | 0.45 (0.43–0.47) | <0.001e |
| Abstinence time (days) | ||||||||
| <2 | 258 | 352 | 73.3 | <0.001 | 2.44 (1.91–3.11) | <0.001f | 1.36 (1.20–1.54) | <0.001f |
| 2–7 | 62 650 | 121 745 | 51.5 | Reference | Reference | Reference | Reference | Reference |
| >7 | 6222 | 9086 | 68.5 | <0.001 | 2.93 (2.79–3.09) | <0.001f | 1.48 (1.44–1.52) | <0.001f |
aP value of univariate analysis; bP value of multivariable Logistic regression; cP value of multivariable Poisson regression; dAdjusted for diagnosis, abstinence time, semen volume, year, and pH value; eAdjusted for diagnosis, age, abstinence time, semen volume, and pH value; fAdjusted for diagnosis, age, semen volume, year, and pH value. RR: risk ratio; OR: odds ratio; -: not calculable in the overall population; CI: confidence interval
Supplementary Table 3.
The effect of Ureaplasma urealyticum on semen quality in the overall population
| Variables | Abnormal | Normal | Total | Abnormal rate (%) | P a | Adjusted OR (95% CI) of logistic regression | P b | Adjusted RR (95% CI) of Poisson regression | P c |
|---|---|---|---|---|---|---|---|---|---|
| Semen volume (N ≥1.4 ml; A <1.4 ml) | |||||||||
| Total | 362 | 10 856 | 11 218 | 3.2 | - | - | - | - | - |
| UU-negative | 293 | 8315 | 8608 | 3.4 | Reference | Reference | Reference | Reference | Reference |
| UU-positive | 69 | 2541 | 2610 | 2.6 | 0.054 | 0.84 (0.64–1.10) | 0.196d | 0.87 (0.69–1.11) | 0.261d |
| pH value (N ≥7.2; A<7.2) | |||||||||
| Total | 2145 | 9073 | 11 218 | 19.1 | - | - | - | - | - |
| UU-negative | 1614 | 6994 | 8608 | 18.8 | Reference | Reference | Reference | Reference | Reference |
| UU-positive | 531 | 2079 | 2610 | 20.3 | 0.07 | 0.96 (0.86–1.08) | 0.525d | 0.97 (0.88–1.07) | 0.593d |
| Total sperm count (N ≥39×106/ejaculate; A <39×106/ejaculate) | |||||||||
| Total | 388 | 10 830 | 11 218 | 3.5 | - | - | - | - | - |
| UU-negative | 297 | 8311 | 8608 | 3.5 | Reference | Reference | Reference | Reference | Reference |
| UU-positive | 91 | 2519 | 2610 | 3.5 | 0.929 | 1.11 (0.87–1.42) | 0.397d | 1.08 (0.88–1.33) | 0.466d |
| Total motility (N ≥42%; A <42%) | |||||||||
| Total | 1017 | 10 201 | 11 218 | 9.1 | - | - | - | - | - |
| UU-negative | 812 | 7796 | 8608 | 9.4 | Reference | Reference | Reference | Reference | Reference |
| UU-positive | 205 | 2405 | 2610 | 79 | 0.014 | 0.98 (0.83–1.16) | 0.833d | 0.99 (0.85–1.14) | 0.841d |
| Sperm vitality (N ≥54%; A <54%) | |||||||||
| Total | 370 | 10 848 | 11 218 | 3.3 | - | - | - | - | - |
| UU-negative | 289 | 8319 | 8608 | 3.4 | Reference | Reference | Reference | Reference | Reference |
| UU-positive | 81 | 2529 | 2610 | 3.1 | 0.525 | 1.04 (0.80–1.34) | 0.783d | 1.02 (0.82–1.28) | 0.841d |
| Sperm concentration (N ≥16×106/ml; A, <16×106/ml) | |||||||||
| Total | 649 | 10 569 | 11 218 | 5.8 | - | - | - | - | - |
| UU-negative | 505 | 8103 | 8608 | 5.9 | Reference | Reference | Reference | Reference | Reference |
| UU-positive | 144 | 2466 | 2610 | 5.5 | 0.503 | 1.08 (0.89–1.32) | 0.416d | 1.06 (0.90–1.26) | 0.491d |
| Normal spermatozoa (stained) (N ≥4%; A <4%) | |||||||||
| Total | 8261 | 2957 | 11 218 | 73.6 | - | - | - | - | - |
| UU-negative | 6411 | 2197 | 8608 | 74.5 | Reference | Reference | Reference | Reference | Reference |
| UU-positive | 1850 | 760 | 2610 | 70.9 | <0.001 | 0.98 (0.89–1.09) | 0.722d | 0.99 (0.90–1.08) | 0.748d |
| Progressive sperm motility (N ≥30%; A <30%) | |||||||||
| Total | 403 | 10 815 | 11 218 | 3.6 | - | - | - | - | - |
| UU-negative | 313 | 8295 | 8608 | 3.6 | Reference | Reference | Reference | Reference | Reference |
| UU-positive | 90 | 2520 | 2610 | 3.5 | 0.651 | 1.12 (0.87–1.43) | 0.373d | 1.08 (0.88–1.34) | 0.457d |
| Teratozoospermia index (N≤1.6; A >1.6) | |||||||||
| Total | 3434 | 7784 | 11 218 | 30.6 | - | - | - | - | - |
| UU-negative | 2542 | 6066 | 8608 | 29.5 | Reference | Reference | Reference | Reference | Reference |
| UU-positive | 892 | 1718 | 2610 | 34.2 | <0.001 | 1.07 (0.97–1.18) | 0.165d | 1.05 (0.97–1.14) | 0.233d |
| Sperm deformity index (N ≤1.6; A >1.6) | |||||||||
| Total | 3194 | 8024 | 11 218 | 28.5 | - | - | - | - | - |
| UU-negative | 2361 | 6247 | 8608 | 27.4 | Reference | Reference | Reference | Reference | Reference |
| UU-positive | 833 | 1777 | 2610 | 31.9 | <0.001 | 1.06 (0.96–1.17) | 0.229d | 1.05 (0.96–1.14) | 0.299d |
aP value of univariate analysis; bP value of multivariable Logistic regression; cP value of multivariable Poisson regression; dAdjusted for diagnosis, age, abstinence time, and year. UU: Ureaplasma urealyticum; RR: risk ratio; OR: odds ratio; -: not calculable; CI: confidence interval
Supplementary Table 4.
The effect of Neisseria gonorrhoeae on semen quality in the overall population
| Variables | Abnormal | Normal | Total | Abnormal rate (%) | P a | Adjusted OR (95% CI) of logistic regression | P b | Adjusted RR (95% CI) of Poisson regression | P c |
|---|---|---|---|---|---|---|---|---|---|
| Semen volume (N ≥1.4 ml; A <1.4 ml) | |||||||||
| Total | 1553 | 31 045 | 32 598 | 4.8 | - | - | - | - | - |
| NG-negative | 1548 | 31 007 | 32 555 | 4.8 | Reference | Reference | Reference | Reference | Reference |
| NG-positive | 5 | 38 | 43 | 11.6 | 0.034 | 2.41 (0.92–6.29) | 0.073d | 2.39 (0.92–6.19) | 0.074d |
| pH value (N ≥7.2; A <7.2) | |||||||||
| Total | 1318 | 31 280 | 32 598 | 4.0 | - | - | - | - | - |
| NG-negative | 1314 | 31 241 | 32 555 | 4.0 | Reference | Reference | Reference | Reference | Reference |
| NG-positive | 4 | 39 | 43 | 9.3 | 0.08 | 1.72 (0.61–4.86) | 0.306d | 1.71 (0.61–4.81) | 0.308d |
| Total sperm count (N ≥39×106/ejaculate; A <39×106/ejaculate) | |||||||||
| Total | 1923 | 30 675 | 32 598 | 5.9 | - | - | - | - | - |
| NG-negative | 1909 | 30 646 | 32 555 | 5.9 | Reference | Reference | Reference | Reference | Reference |
| NG-positive | 14 | 29 | 43 | 32.7 | <0.001 | 7.24 (3.77–13.90) | <0.001d | 7.03 (3.69–13.40) | <0.001d |
| Total motility (N ≥42%; A <42%) | |||||||||
| Total | 5306 | 27 292 | 32 598 | 16.3 | - | - | - | - | - |
| NG-negative | 5289 | 27 266 | 32 555 | 16.3 | Reference | Reference | Reference | Reference | Reference |
| NG-positive | 17 | 26 | 43 | 39.5 | <0.001 | 3.11 (1.67–5.79) | <0.001d | 3.07 (1.65–5.70) | <0.001d |
| Sperm vitality (N ≥54%; A <54%) | |||||||||
| Total | 2070 | 30 528 | 32 598 | 6.4 | - | - | - | - | - |
| NG-negative | 2055 | 30 500 | 32 555 | 6.3 | Reference | Reference | Reference | Reference | Reference |
| NG-positive | 15 | 28 | 43 | 34.9 | <0.001 | 6.02 (3.14–11.53) | <0.001d | 5.84 (3.06–11.14) | <0.001d |
| Sperm concentration (N ≥16×106/ml; A <16×106/ml) | |||||||||
| Total | 3996 | 28 602 | 32 598 | 12.3 | - | - | - | - | - |
| NG-negative | 3982 | 28 573 | 32 555 | 12.2 | Reference | Reference | Reference | Reference | Reference |
| NG-positive | 14 | 29 | 43 | 32.6 | <0.001 | 3.68 (1.92–7.07) | <0.001d | 3.63 (1.90–6.94) | <0.001d |
| Normal spermatozoa (stained) (N ≥4%; A <4%) | |||||||||
| Total | 12 378 | 20 220 | 32 598 | 38.0 | - | - | - | - | - |
| NG-negative | 12 359 | 20 196 | 32 555 | 38.0 | Reference | Reference | Reference | Reference | Reference |
| NG-positive | 19 | 24 | 43 | 44.2 | 0.401 | 0.87 (0.32–2.38) | 0.792d | 0.87 (0.32–2.37) | 0.792d |
| Progressive sperm motility (N ≥30%; A <30%) | |||||||||
| Total | 2594 | 30 004 | 32 598 | 8.0 | - | - | - | - | - |
| NG-negative | 2578 | 29 977 | 32 555 | 7.9 | Reference | Reference | Reference | Reference | Reference |
| NG-positive | 16 | 27 | 43 | 37.2 | <0.001 | 5.07 (2.66–9.65) | <0.001d | 4.95 (2.61–9.38) | <0.001d |
| Teratozoospermia index (N ≤1.6; A >1.6) | |||||||||
| Total | 20 561 | 12 037 | 32 598 | 63.1 | - | - | - | - | - |
| NG-negative | 20 536 | 12 019 | 32 555 | 63.1 | Reference | Reference | Reference | Reference | Reference |
| NG-positive | 25 | 18 | 43 | 58.1 | 0.502 | 1.32 (0.51–3.40) | 0.567d | 1.32 (0.51–3.39) | 0.568d |
| Sperm deformity index (N ≤1.6; A >1.6) | |||||||||
| Total | 20 417 | 12 181 | 32 598 | 62.6 | - | - | - | - | - |
| NG-negative | 20 392 | 12 163 | 32 555 | 62.6 | Reference | Reference | Reference | Reference | Reference |
| NG-positive | 25 | 18 | 43 | 58.1 | 0.542 | 1.38 (0.53–3.57) | 0.508d | 1.38 (0.53–3.55) | 0.509d |
aP value of univariate analysis; bP value of multivariable Logistic regression; cP value of multivariable Poisson regression; dAdjusted for diagnosis, age, abstinence time, and year. NG: Neisseria gonorrhoeae; RR: risk ratio; OR: odds ratio; -: not calculable; CI: Confidence interval
Supplementary Table 5.
The effect of Mycoplasma genitalium on semen quality in the overall population
| Variables | Abnormal | Normal | Total | Abnormal rate (%) | P a | Adjusted OR (95% CI) of logistic regression | P b | Adjusted RR (95% CI) of Poisson regression | P c |
|---|---|---|---|---|---|---|---|---|---|
| Semen volume (N ≥1.4 ml; A <1.4 ml) | |||||||||
| Total | 1454 | 29 234 | 30 688 | 4.7 | - | - | - | - | - |
| MG-negative | 1423 | 28 801 | 30 224 | 4.7 | Reference | Reference | Reference | Reference | Reference |
| MG-positive | 31 | 433 | 464 | 6.7 | 0.047 | 1.37 (0.941.98) | 0.097d | 1.36 (0.94–1.96) | 0.100d |
| pH value (N ≥7.2; A <7.2) | |||||||||
| Total | 1350 | 29 338 | 30 688 | 4.4 | - | - | - | - | - |
| MG-negative | 1316 | 28 908 | 30 224 | 4.4 | Reference | Reference | Reference | Reference | Reference |
| MG-positive | 34 | 430 | 464 | 7.3 | 0.002 | 1.68 (1.18–2.41) | 0.004d | 1.66 (1.17–2.37) | 0.004d |
| Total sperm count (N≥39×106/ejaculate; A <39×106/ejaculate) | |||||||||
| Total | 1797 | 28 891 | 30 688 | 5.9 | - | - | - | - | - |
| MG-negative | 1758 | 28 466 | 30 224 | 5.8 | Reference | Reference | Reference | Reference | Reference |
| MG-positive | 39 | 425 | 464 | 8.4 | 0.018 | 1.45 (1.04–2.03) | 0.027d | 1.44 (1.04–2.00) | 0.029d |
| Total motility (N ≥42%; A <42%) | |||||||||
| Total | 4955 | 25 733 | 30 688 | 16.2 | - | - | - | - | - |
| MG-negative | 4888 | 25 336 | 30 224 | 16.2 | Reference | Reference | Reference | Reference | Reference |
| MG-positive | 67 | 397 | 464 | 14.4 | 0.314 | 0.80 (0.61–1.04) | 0.094d | 0.80 (0.62–1.04) | 0.097d |
| Sperm vitality (N ≥54%; A <54%) | |||||||||
| Total | 1992 | 28 696 | 30 688 | 6.5 | - | - | - | - | - |
| MG-negative | 1969 | 28 255 | 30 224 | 6.5 | Reference | Reference | Reference | Reference | Reference |
| MG-positive | 23 | 441 | 464 | 5.0 | 0.176 | 0.72 (0.47–1.10) | 0.126d | 0.72 (0.47–1.10) | 0.129d |
| Sperm concentration (N ≥16×106/ml; A <16×106/ml) | |||||||||
| Total | 3756 | 26 932 | 30 688 | 12.2 | - | - | - | - | - |
| MG-negative | 3672 | 26 552 | 30 224 | 12.2 | Reference | Reference | Reference | Reference | Reference |
| MG-positive | 84 | 380 | 464 | 18.1 | <0.001 | 1.57 (1.24–2.00) | <0.001d | 1.56 (1.23–1.98) | <0.001d |
| Normal spermatozoa (stained) (N ≥4%; A <4%) | |||||||||
| Total | 10 775 | 19 913 | 30 688 | 35.1 | - | - | - | - | - |
| MG-negative | 10 589 | 19 635 | 30 224 | 35.0 | Reference | Reference | Reference | Reference | Reference |
| MG-positive | 186 | 278 | 464 | 40.1 | 0.024 | 0.84 (0.62–1.13) | 0.242d | 0.84 (0.62–1.13) | 0.247d |
| Progressive sperm motility (N ≥30%; A <30%) | |||||||||
| Total | 2509 | 28 179 | 30 688 | 8.2 | - | - | - | - | - |
| MG-negative | 2478 | 27 746 | 30 224 | 8.2 | Reference | Reference | Reference | Reference | Reference |
| MG-positive | 31 | 433 | 464 | 6.7 | 0.236 | 0.78 (0.54–1.14) | 0.199d | 0.79 (0.55–1.14) | 0.203d |
| Teratozoospermia index (N ≤1.6; A >1.6) | |||||||||
| Total | 20 161 | 10 527 | 30 688 | 65.7 | - | - | - | - | - |
| MG-negative | 19 878 | 10 346 | 30 224 | 65.8 | Reference | Reference | Reference | Reference | Reference |
| MG-positive | 283 | 181 | 464 | 61.0 | 0.031 | 1.19 (0.89–1.59) | 0.242d | 1.19 (0.89–1.58) | 0.248d |
| Sperm deformity index (N ≤1.6; A >1.6) | |||||||||
| Total | 20 060 | 10 628 | 30 688 | 65.4 | - | - | - | - | - |
| MG-negative | 19 780 | 10 444 | 30 224 | 65.4 | Reference | Reference | Reference | Reference | Reference |
| MG-positive | 280 | 184 | 464 | 60.3 | 0.022 | 1.17 (0.87–1.57) | 0.300d | 1.17 (0.87–1.56) | 0.305d |
aP value of univariate analysis; bP value of multivariable Logistic regression; cP value of multivariable Poisson regression; dAdjusted for diagnosis, age, abstinence time, and year. MG: Mycoplasma genitalium; RR: risk ratio; OR: odds ratio; -: not calculable; CI: confidence interval
Supplementary Table 6.
The effect of Antisperm antibody on semen quality in the overall population
| Variables | Abnormal | Normal | Total | Abnormal rate (%) | P a | Adjusted OR (95% CI) of logistic regression | P b | Adjusted RR (95% CI) of Poisson regression | P c |
|---|---|---|---|---|---|---|---|---|---|
| Semen volume (N ≥1.4 ml; A <1.4 ml) | |||||||||
| Total | 3093 | 78 177 | 81 270 | 3.8 | - | - | - | - | - |
| AsAb-negative | 3032 | 76 598 | 79 630 | 3.8 | Reference | Reference | Reference | Reference | Reference |
| AsAb-positive | 61 | 1579 | 1640 | 3.7 | 0.854 | 0.85 (0.66–1.10) | 0.223d | 0.85 (0.66–1.10) | 0.230d |
| pH value (N ≥7.2; A <7.2) | |||||||||
| Total | 9254 | 72 016 | 81 270 | 11.4 | - | - | - | - | - |
| AsAb-negative | 9205 | 70 425 | 79 630 | 11.6 | Reference | Reference | Reference | Reference | Reference |
| AsAb-positive | 49 | 1591 | 1640 | 3.0 | <0.001 | 0.28 (0.21–0.38) | <0.001d | 0.29 (0.22–0.38) | <0.001d |
| Total sperm count (N ≥39×106/ejaculate; A <39×106/ejaculate) | |||||||||
| Total | 4600 | 76 670 | 81 270 | 5.7 | - | - | - | - | - |
| AsAb-negative | 4487 | 75 143 | 79 630 | 5.6 | Reference | Reference | Reference | Reference | Reference |
| AsAb-positive | 113 | 1527 | 1640 | 6.9 | 0.029 | 1.16 (0.96–1.41) | 0.128d | 1.16 (0.96–1.40) | 0.133d |
| Total motility (N ≥42%; A <42%) | |||||||||
| Total | 10 812 | 70 458 | 81 270 | 13.3 | - | - | - | - | - |
| AsAb-negative | 10 434 | 69 196 | 79 630 | 13.1 | Reference | Reference | Reference | Reference | Reference |
| AsAb-positive | 378 | 1262 | 1640 | 23.1 | <0.001 | 1.89 (1.68–2.13) | <0.001d | 1.85 (1.65–2.09) | <0.001d |
| Sperm vitality (N ≥54%; A <54%) | |||||||||
| Total | 4266 | 77 004 | 81 270 | 5.3 | - | - | - | - | - |
| AsAb-negative | 4119 | 75 511 | 79 630 | 5.2 | Reference | Reference | Reference | Reference | Reference |
| AsAb-positive | 147 | 1493 | 1640 | 9.0 | <0.001 | 1.56 (1.31–1.86) | <0.001d | 1.54 (1.30–1.83) | <0.001d |
| Sperm concentration (N ≥16×106/ml; A <16×106/ml) | |||||||||
| Total | 6493 | 74 777 | 81 270 | 8.0 | - | - | - | - | - |
| AsAb-negative | 6344 | 73 286 | 79 630 | 8.0 | Reference | Reference | Reference | Reference | Reference |
| AsAb-positive | 149 | 1491 | 1640 | 9.1 | 0.098 | 1.12 (0.94–1.33) | 0.201d | 1.12 (0.94–1.32) | 0.206d |
| Normal spermatozoa (stained) (N ≥4%; A <4%) | |||||||||
| Total | 55 278 | 25 992 | 81 270 | 68.0 | - | - | - | - | - |
| AsAb-negative | 54 746 | 24 884 | 79 630 | 68.8 | Reference | Reference | Reference | Reference | Reference |
| AsAb-positive | 532 | 1108 | 1640 | 32.4 | <0.001 | 0.22 (0.19–0.25) | <0.001d | 0.23 (0.20–0.26) | <0.001d |
| Progressive sperm motility (N ≥30%; A <30%) | |||||||||
| Total | 5222 | 76 048 | 81 270 | 6.4 | - | - | - | - | - |
| AsAb-negative | 5015 | 74 615 | 79 630 | 6.3 | Reference | Reference | Reference | Reference | Reference |
| AsAb-positive | 207 | 1433 | 1640 | 12.6 | <0.001 | 1.80 (1.54–2.09) | <0.001d | 1.76 (1.52–2.04) | <0.001d |
| Teratozoospermia index (N ≤1.6; A >1.6) | |||||||||
| Total | 28 694 | 52 576 | 81 270 | 35.3 | - | - | - | - | - |
| AsAb-negative | 27 571 | 52 059 | 79 630 | 34.6 | Reference | Reference | Reference | Reference | Reference |
| AsAb-positive | 1123 | 517 | 1640 | 68.5 | <0.001 | 3.56 (3.15–4.01) | <0.001d | 3.45 (3.07–3.89) | <0.001d |
| Sperm deformity index (N ≤1.6; A >1.6) | |||||||||
| Total | 27 700 | 53 570 | 81 270 | 34.1 | - | - | - | - | - |
| AsAb-negative | 26 581 | 53 049 | 79 630 | 33.4 | Reference | Reference | Reference | Reference | Reference |
| AsAb-positive | 1119 | 521 | 1640 | 68.2 | <0.001 | 3.87 (3.42–4.37) | <0.001d | 3.75 (3.32–4.23) | <0.001d |
aP value of univariate analysis; bP value of multivariable Logistic regression; cP value of multivariable Poisson regression; dAdjusted for diagnosis, age, abstinence time, and year. AsAb: antisperm antibody; RR: risk ratio; OR: odds ratio; -: not calculable; CI: confidence interval
We further investigated the impact of reproductive tract pathogens on semen quality in infertile males. The results revealed that these pathogens exerted significant adverse effects on semen parameters. Specifically, patients positive for CT showed significantly lower levels of pH value, total motility, sperm vitality, normal sperm (stained), and progressive sperm motility than CT-negative patients. Similarly, compared with UU-negative patients, UU-positive patients exhibited reduced sperm vitality, sperm concentration, and progressive sperm motility. Moreover, compared with MG-negative patients, MG-positive patients had markedly lower total sperm counts and sperm concentrations. NG infection demonstrated the most pronounced effect on semen quality; NG-positive patients exhibited more than a threefold reduction in total sperm count, total motility, sperm vitality, sperm concentration, and progressive sperm motility rate than NG-negative patients (all P < 0.05; Table 3).
Table 4.
The impact of the clearance of Neisseria gonorrhoeae, Chlamydia trachomatis, Ureaplasma urealyticum, and Mycoplasma genitalium on pregnancy rates in infertile men
| Pathogen | Turned negative | Not turned negative | OR | Lower limit | Upper limit | P | ||
|---|---|---|---|---|---|---|---|---|
|
|
|
|||||||
| + | − | + | − | |||||
| CT | 163 | 51 | 271 | 1449 | 17.09 | 12.16 | 24.01 | <0.001 |
| UU | 423 | 108 | 1245 | 609 | 1.92 | 1.52 | 2.42 | <0.001 |
| NG | 4 | 7 | 11 | 33 | 1.71 | 0.42 | 6.99 | 0.468 |
| MG | 78 | 225 | 216 | 477 | 0.77 | 0.57 | 1.04 | 0.097 |
+: pregnancy; −: unpregnancy; OR: odds ratio; CT: Chlamydia trachomatis; UU: Ureaplasma urealyticum; NG: Neisseria gonorrhoeae; MG: Mycoplasma genitalium
Table 3.
The impact of genital tract pathogens on semen quality in infertile men
| Semen factor | Negative/total (%) | Positive/total (%) | OR (95% CI) | P |
|---|---|---|---|---|
| CT | ||||
| Semen volume | 934/18 147 (5.1) | 90/1594 (5.6) | 1.11 (0.89–1.38) | 0.422 |
| pH | 1299/18 147 (7.2) | 151/1594 (9.5) | 1.36 (1.14–1.62) | 0.001 |
| Total sperm count | 1128/18 147 (6.2) | 110/1594 (6.9) | 1.12 (0.92–1.37) | 0.304 |
| Total motility | 2574/18 147 (14.2) | 274/1594 (17.2) | 1.26 (1.1–1.44) | 0.001 |
| Sperm vitality | 1102/18 147 (6.1) | 145/1594 (9.1) | 1.55 (1.29–1.86) | <0.001 |
| Sperm concentration | 2435/18 147 (13.4) | 239/1594 (15.0) | 1.14 (0.99–1.32) | 0.085 |
| Normal sperm (stained) | 7297/18 147 (40.2) | 765/1594 (48.0) | 1.37 (1.24–1.52) | <0.001 |
| Progressive sperm motility | 1294/18 147 (7.1) | 189/1594 (11.9) | 1.76 (1.49–2.06) | <0.001 |
| Teratozoospermia index | 9665/18 147 (53.3) | 857/1594 (53.8) | 1.02 (0.92–1.13) | 0.718 |
| Sperm deformity index | 9576/18 147 (52.8) | 852/1594 (53.5) | 1.03 (0.93–1.14) | 0.620 |
| UU | ||||
| Semen volume | 33/1188 (2.8) | 17/360 (4.7) | 1.76 (0.97–3.17) | 0.097 |
| pH | 122/1188 (10.3) | 43/360 (11.9) | 1.19 (0.83–1.72) | 0.421 |
| Total sperm count | 62/1188 (5.2) | 28/360 (7.8) | 1.54 (0.98–2.45) | 0.091 |
| Total motility | 182/1188 (15.3) | 68/360 (18.9) | 1.29 (0.95–1.76) | 0.126 |
| Sperm vitality | 61/1188 (5.1) | 31/360 (8.6) | 1.75 (1.12–2.74) | 0.021 |
| Sperm concentration | 104/1188 (8.8) | 47/360 (13.1) | 1.57 (1.09–2.27) | 0.021 |
| Normal sperm (stained) | 1036/1188 (87.2) | 310/360 (86.1) | 0.9 (0.64–1.27) | 0.652 |
| Progressive sperm motility | 66/1188 (5.6) | 35/360 (9.7) | 1.84 (1.2–2.82) | 0.007 |
| Teratozoospermia index | 230/1188 (19.4) | 82/360 (22.8) | 1.23 (0.93–1.64) | 0.180 |
| Sperm deformity index | 190/1188 (16.0) | 71/360 (19.7) | 1.29 (0.96–1.75) | 0.115 |
| MG | ||||
| Semen volume | 761/13 967 (5.4) | 18/234 (7.7) | 1.48 (0.92–2.4) | 0.177 |
| pH | 695/13 967 (5.0) | 14/234 (6.0) | 1.25 (0.73–2.15) | 0.582 |
| Total sperm count | 864/13 967 (6.2) | 24/234 (10.3) | 1.76 (1.15–2.7) | 0.016 |
| Total motility | 2195/13 967 (15.7) | 34/234 (14.5) | 0.92 (0.64–1.33) | 0.686 |
| Sperm vitality | 949/13 967 (6.8) | 14/234 (6.0) | 0.9 (0.53–1.54) | 0.720 |
| Sperm concentration | 2063/13 967 (14.8) | 59/234 (25.2) | 1.96 (1.45–2.63) | <0.001 |
| Normal sperm (stained) | 4173/13 967 (29.9) | 65/234 (27.8) | 0.91 (0.68–1.21) | 0.533 |
| Progressive sperm motility | 1181/13 967 (8.5) | 20/234 (8.5) | 1.03 (0.65–1.63) | 1.000 |
| Teratozoospermia index | 9907/13 967 (70.9) | 170/234 (72.6) | 1.08 (0.81–1.45) | 0.616 |
| Sperm deformity index | 9866/13 967 (70.6) | 170/234 (72.6) | 1.1 (0.82–1.47) | 0.550 |
| NG | ||||
| Semen volume | 800/15 064 (5.3) | 4/32 (12.5) | 2.81 (1.04–7.62) | 0.088 |
| pH | 750/15 064 (5.0) | 3/32 (9.4) | 2.26 (0.75–6.87) | 0.213 |
| Total sperm count | 933/15 064 (6.2) | 11/32 (34.4) | 8.1 (3.94–16.62) | <0.001 |
| Total motility | 2371/15 064 (15.7) | 16/32 (50.0) | 5.35 (2.7–10.61) | <0.001 |
| Sperm vitality | 990/15 064 (6.6) | 14/32 (43.8) | 11.14 (5.58–22.21) | <0.001 |
| Sperm concentration | 2158/15 064 (14.3) | 12/32 (37.5) | 3.65 (1.8–7.38) | 0.001 |
| Normal sperm (stained) | 5099/15 064 (33.8) | 14/32 (43.8) | 1.53 (0.77–3.05) | 0.320 |
| Progressive sperm motility | 1234/15 064 (8.2) | 15/32 (46.9) | 9.92 (5–19.71) | <0.001 |
| Teratozoospermia index | 10 130/15 064 (67.2) | 19/32 (59.4) | 0.7 (0.35–1.41) | 0.448 |
| Sperm deformity index | 10 060/15 064 (66.8) | 19/32 (59.4) | 0.72 (0.36–1.44) | 0.483 |
| Herpes simplex virus II | ||||
| Semen volume | 1/14 (7.1) | - | - | - |
| pH | 2/14 (14.3) | - | - | - |
| Total sperm count | 2/14 (14.3) | - | - | - |
| Total motility | 2/14 (14.3) | - | - | - |
| Sperm vitality | 1/14 (7.1) | - | - | - |
| Sperm concentration | 1/14 (7.1) | - | - | - |
| Normal sperm (stained) | 3/14 (21.4) | - | - | - |
| Progressive sperm motility | 1/14 (7.1) | - | - | - |
| Teratozoospermia index | 11/14 (78.6) | - | - | - |
| Sperm deformity index | 11/14 (78.6) | - | - | - |
OR: odds ratio; -: not calculable; CI: confidence interval; CT: Chlamydia trachomatis; UU: Ureaplasma urealyticum; NG: Neisseria gonorrhoeae; MG: Mycoplasma genitalium
Effect of pathogen clearance on pregnancy outcomes
To further evaluate clinical outcomes, data of the infertile men who initially tested positive for NG, CT, UU, or MG and whose follow-up data were available were analyzed. The study included 1934 patients with CT, 2385 with UU, 55 with NG, and 996 with MG. Clearance of CT and UU was significantly associated with improved pregnancy rates. Infertile men who cleared CT achieved pregnancy 17.09 times more often than those with persistent infection (OR = 17.09, 95% CI: 12.16–24.01; P < 0.001). Similarly, clearance of UU increased pregnancy rates nearly twofold (OR = 1.92, 95% CI: 1.52–2.42; P < 0.001). In contrast, the clearance of NG and MG did not significantly affect pregnancy outcomes (both P > 0.05; Figure 1b and Table 4).
DISCUSSION
In recent years, the prevalence of infertility has increased markedly, posing not only a substantial psychological and economic burden on affected couples but also a significant public health challenge.19 Infertility contributes to emotional stress, marital discord, social isolation, and financial strain, representing a considerable societal impact.10 Approximately 15% of couples of reproductive age worldwide experience infertility.20 In China, the prevalence of infertility among reproductive-age couples is even higher, with male factors contributing to 40%–50% of cases.21 Etiologies of male infertility include genitourinary infections, endocrine disorders, abnormal reproductive organ development, genetic abnormalities, and micronutrient deficiencies.11 Semen parameters remain critical indicators of male reproductive function and provide essential guidance for clinical evaluation and management.22
In contrast to previous studies,7,8,10,16,23 our research provides several novel insights into male infertility. First, this work represents one of the large population-based analyses of infertile men in China, encompassing more than 130 000 participants over an 8-year period. The large-scale design not only enhances statistical reliability but also improves the generalizability of the findings. Second, unlike earlier studies that focused on either age-related changes or pathogen-specific effects in isolation, our study simultaneously evaluated the combined and independent effects of age, genital infections, and AsAb on semen quality and infertility risk. This integrative approach offers a more comprehensive understanding of multifactorial influences on male reproductive health. Third, we identified a U-shaped association between age and infertility risk, with the lowest risk observed among men aged 25–29 years. This nonlinear pattern has not been reported previously in Chinese populations and highlights potential reproductive risks at both younger and older ages. Fourth, by concurrently assessing multiple pathogens, our analysis revealed that CT has the strongest detrimental effects on semen quality and fertility. Finally, our findings demonstrate that the clearance of CT and UU infections significantly improves pregnancy outcomes, providing direct clinical evidence that early detection and targeted treatment of genital infections can increase reproductive success. Collectively, these findings expand the current understanding of male infertility and offer valuable implications for both clinical management and public health interventions.
Immune-mediated infertility accounts for approximately 15%–30% of male infertility cases and is often associated with AsAb.24 Under physiological conditions, the blood–testis and blood–epididymis barriers prevent AsAb production.17 However, conditions such as epididymitis, testicular trauma, vas deferens obstruction, or testicular inflammation can expose sperm antigens to the immune system, triggering AsAb production.17 The effects of AsAb on sperm function depend on the binding site: binding to the sperm head can disrupt acrosomal enzyme activity and nuclear integrity, impairing sperm–egg fusion and embryonic development; binding to the tail reduces motility by altering flagellar movement; and binding to the midpiece may affect mitochondrial DNA, inducing apoptosis.15,16,17 In our cohort, infertile men exhibited significantly higher AsAb positivity than controls. AsAb presence was associated with reduced sperm vitality, total motility, progressive motility, and normal sperm counts, as well as increased teratozoospermia and sperm deformity indices, confirming the negative impact of immune-mediated factors on semen quality.
Urogenital tract infections are a major contributor to male infertility, accounting for 10%–40% of cases.12 In China, Mycoplasma and Chlamydia infections contribute to 20%–40% of nongonococcal urethritis and 35%–50% of male genital tract infections, respectively.1 CT, a common sexually transmitted pathogen, induces genital tract inflammation that may progress to prostatitis or epididymitis via host immune activation.12,25 Infection can increase sperm apoptosis by disrupting phosphatidylserine exposure and DNA integrity, ultimately impairing sperm motility.26 In our study, CT positivity was significantly greater among infertile men and was associated with reduced sperm vitality, decreased normal sperm counts, and elevated teratozoospermia and sperm deformity indices.
NG predominantly affects young adult males, and epididymo-orchitis secondary to infection can result in persistent oligozoospermia or azoospermia.25 In this cohort, NC infection was associated with decreased total sperm count, motility, vitality, concentration, and progressive motility. Mechanistically, gonococcal fimbriae may adhere to the seminal vesicles and prostate, triggering inflammation that impairs spermatogenesis and sperm motility.8 Latent infections in untreated individuals may facilitate pathogen transmission, highlighting both reproductive and public health implications.
Mycoplasma species, including UU and MG, are opportunistic pathogens capable of directly invading spermatogenic cells and disrupting sperm development.10 Their protoplasmic membrane can fuse with sperm membranes, allowing cytoplasmic proteins and lipids to interfere with sperm production, leading to decreased sperm counts and motility.11,14 High Mycoplasma loads can cause sperm aggregation and morphological distortion, further impairing motility.25,27 In our study, univariate analysis suggested that UU infection adversely affected semen parameters; however, multivariate analysis adjusted for age and testing year attenuated these associations. In contrast, MG infection was consistently associated with reduced semen quality. These findings indicate that while Mycoplasma infections can negatively affect spermatozoa, their impact may vary depending on pathogen load, coinfections, and host factors, warranting further investigation.
Despite the large sample size and comprehensive analysis, this study has several limitations. First, pregnancy outcomes for partners were unavailable for all participants, limiting our ability to directly correlate semen parameter values and microbial infections with fertility outcomes. Second, as a single-center study, our findings may not be fully generalizable to other regions, although the large sample size and population coverage in Southwest China increase reliability. Third, only five specific microorganisms were analyzed; the potential influence of other pathogens and the broader seminal microbiome remain to be explored. Future multicenter studies incorporating comprehensive microbiological profiling are needed to elucidate better pathogen-mediated mechanisms in male infertility.
In conclusion, semen quality is significantly lowered in infertile men than that in healthy controls, with a U-shaped age-related risk profile. Immune factors (AsAb) and genital pathogens, including CT, NG, UU, and MG, are associated with impaired semen quality and increased infertility risk. These findings underscore the importance of early detection and management of urogenital infections and immune factors in men who present with infertility and provide insights for targeted clinical interventions and future mechanistic studies.
AUTHOR CONTRIBUTIONS
JJC conceptualized the study, visualized the data, developed the methodology used, applied the software, performed the statistical analysis, wrote the original draft, and reviewed and edited the manuscript. WZ applied the analyses with software, performed the statistical analysis, and developed the methodology used. YQZ and XYZ visualized the data. SL applied the analyses with software, performed the statistical analysis, and reviewed the manuscript. HYZ visualized the data and developed the methodology. CLL conceptualized the study and wrote, reviewed, and edited the manuscript. JJC and CLL confirm the authenticity of all the raw data. All authors read and approved the final manuscript.
COMPETING INTERESTS
All authors declare no competing interests.
Supplementary Information is linked to the online version of the paper on the Asian Journal of Andrology website.
Flowchart of patient inclusion. AsAb: antisperm antibodies; NG: Neisseria gonorrhea; CT: Chlamydia trachomatis; UU: Ureaplasma urealyticum; MG: Mycoplasma genitalium; HSV-II: herpes simplex virus II.
Number of semen analyses performed and the incidence of infertility in different age groups. (a) Number of semen analyses. (b) Incidence of male infertility.
REFERENCES
- 1.He Z, Xie M, Li QQ, Duan J, Lu X. Research progress on the microregulatory mechanisms of fertilization: a review. In Vivo. 2022;36:2002–13. doi: 10.21873/invivo.12926. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 2.Zegers-Hochschild F, Adamson GD, Dyer S, Racowsky C, de Mouzon J, et al. The international glossary on infertility and fertility care, 2017. Fertil Steril. 2017;108:393–406. doi: 10.1016/j.fertnstert.2017.06.005. [DOI] [PubMed] [Google Scholar]
- 3.Mehta A. Qualitative research in male infertility. Urol Clin North Am. 2020;47:205–10. doi: 10.1016/j.ucl.2019.12.007. [DOI] [PubMed] [Google Scholar]
- 4.Horta F, Vollenhoven B, Healey M, Busija L, Catt S, et al. Male ageing is negatively associated with the chance of live birth in IVF/ICSI cycles for idiopathic infertility. Hum Reprod. 2019;34:2523–32. doi: 10.1093/humrep/dez223. [DOI] [PubMed] [Google Scholar]
- 5.Kaarouch I, Bouamoud N, Madkour A, Louanjli N, Saadani B, et al. Paternal age: negative impact on sperm genome decays and IVF outcomes after 40 years. Mol Reprod Dev. 2018;85:271–80. doi: 10.1002/mrd.22963. [DOI] [PubMed] [Google Scholar]
- 6.Szczygiel M, Kurpisz M. Teratozoospermia and its effect on male fertility potential. Andrologia. 1999;31:63–75. [PubMed] [Google Scholar]
- 7.Yue CB, Zheng XL, Shang W, Wang P, Xie ZH, et al. Assessing intentions for a third child among Chinese adults: a nationwide online survey regarding China's 3-child policy. Medicine (Baltimore) 2023;102:e33255. doi: 10.1097/MD.0000000000033255. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8.Gdoura R, Kchaou W, Ammar-Keskes L, Chakroun N, Sellemi A, et al. Assessment of Chlamydia trachomatis, Ureaplasma urealyticum, Ureaplasma parvum, Mycoplasma hominis, and Mycoplasma genitalium in semen and first void urine specimens of asymptomatic male partners of infertile couples. J Androl. 2008;29:198–206. doi: 10.2164/jandrol.107.003566. [DOI] [PubMed] [Google Scholar]
- 9.Jordan SJ, Toh E, Williams JA, Fortenberry L, LaPradd ML, et al. Aetiology and prevalence of mixed-infections and mono-infections in non-gonococcal urethritis in men: a case-control study. Sex Transm Infect. 2020;96:306–11. doi: 10.1136/sextrans-2019-054121. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 10.Liu A, Zhang L. [Effects of Ureaplasma urealyticum infection on semen quality and serum sex hormone levels in infertile males] Inter Med Heal Guid News. 2024;30:1532–4. [Article in Chinese] [Google Scholar]
- 11.Paira DA, Olivera C, Tissera AD, Molina RI, Olmedo JJ, et al. Ureaplasma urealyticum and Mycoplasma hominis urogenital infections associate with semen inflammation and decreased sperm quality. J Leukoc Biol. 2023;113:18–26. doi: 10.1093/jleuko/qiac006. [DOI] [PubMed] [Google Scholar]
- 12.Keikha M, Hosseininasab-Nodoushan SA, Sahebkar A. Association between Chlamydia trachomatis infection and male infertility: a systematic review and meta-analysis. Mini Rev Med Chem. 2023;23:746–55. doi: 10.2174/1389557522666220827160659. [DOI] [PubMed] [Google Scholar]
- 13.Gdoura R, Kchaou W, Chaari C, Znazen A, Keskes L, et al. Ureaplasma urealyticum, Ureaplasma parvum, Mycoplasma hominis and Mycoplasma genitalium infections and semen quality of infertile men. BMC Infect Dis. 2007;7:129. doi: 10.1186/1471-2334-7-129. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 14.Liang L, Sun B, Cao J, Dong Y, Liu Z. [The effects of male genital tract Ureaplasma urealyticum positive on sperm quality, sperm function and the outcome of assisted reproductive technology] Chinese J Androl. 2021;35:37–41. [Article in Chinese] [Google Scholar]
- 15.Vickram AS, Dhama K, Chakraborty S, Samad HA, Latheef SK, et al. Role of antisperm antibodies in infertility, pregnancy, and potential for contraceptive and antifertility vaccine designs: research progress and pioneering vision. Vaccines. 2019;7:116. doi: 10.3390/vaccines7030116. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 16.Hu YY, Wang LY, Song BT, Cao SS, Chen AL. [Impacts of different procedures of testicular sperm retrieval on testicular function and antisperm antibodies in azoospermia patients] Natl J Androl. 2017;23:620–5. [Article in Chinese] [PubMed] [Google Scholar]
- 17.Zhao Y, Zhao E, Zhang C, Zhang H. Study of the changes of acrosomal enzyme, nitric oxide synthase, and superoxide dismutase of infertile patients with positive antisperm antibody in seminal plasma. Cell Biochem Biophys. 2015;73:639–42. doi: 10.1007/s12013-015-0641-5. [DOI] [PubMed] [Google Scholar]
- 18.World Health Organization. WHO Laboratory Manual for the Examination and Processing of Human Semen. sixth edition. Geneva: World Health Organization; 2021. [Google Scholar]
- 19.Zegers-Hochschild F, Adamson GD, Dyer S, Racowsky C, de Mouzon J, et al. The international glossary on infertility and fertility care, 2017. Hum Reprod. 2017;32:1786–801. doi: 10.1093/humrep/dex234. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 20.Wang Q, Que C, Chen G. Effects of leukocyte elastase in semen on sperm quality. Medicine. 2022;101:e31111. doi: 10.1097/MD.0000000000031111. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 21.Zhou Z, Zheng D, Wu H, Li R, Xu S, et al. Epidemiology of infertility in China: a population-based study. BJOG. 2018;125:432–41. doi: 10.1111/1471-0528.14966. [DOI] [PubMed] [Google Scholar]
- 22.Gao J, Yuan R, Yang S, Wang Y, Huang Y, et al. Age-related changes in human conventional semen parameters and sperm chromatin structure assay-defined sperm DNA/chromatin integrity. Reprod Biomed Online. 2021;42:973–82. doi: 10.1016/j.rbmo.2021.02.006. [DOI] [PubMed] [Google Scholar]
- 23.Zhu JD, Yu Y, Li Y, Zhou LM, Shang XJ. [Correlation of seminal plasma elastase with sperm reactive oxygen species and semen parameters in infertile men] Natl J Androl. 2022;28:223–6. [Article in Chinese] [PubMed] [Google Scholar]
- 24.Zhang H, Zhao E, Zhang C, Li X. The change of semen superoxide dismutase and acrosin activity in the sterility of male patients with positive antisperm antibody. Cell Biochem Biophys. 2015;73:451–3. doi: 10.1007/s12013-015-0663-z. [DOI] [PubMed] [Google Scholar]
- 25.Bao HQ, Sun L, Yang XN, Ding J, Zhang TF. Chlamydia trachomatis (CT), Ureaplasma urealyticum (UU), Neisseria gonorrhoeae (NG) and Mycoplasma genitalium (MG) in male from Chongqing. Chin J Androl. 2020;34:38–41. [Article in Chinese] [Google Scholar]
- 26.Xia Q, Wei R, Cui Y. [Distribution characteristics of pathogens and its influence on semen quality parameters in patients with male infertility. Chin J Gen Pract. 2017;15:759–62. [Article in Chinese] [Google Scholar]
- 27.Zhang HY, Lu WH, Zhao Y, Zhen Y, Gao R, et al. [Clinical observation on impacts of Ureaplasma urealytieum infection upon quality of sperm] J Kunming Med Univ. 2020;41:134–7. [Article in Chinese] [Google Scholar]
Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Supplementary Materials
Flowchart of patient inclusion. AsAb: antisperm antibodies; NG: Neisseria gonorrhea; CT: Chlamydia trachomatis; UU: Ureaplasma urealyticum; MG: Mycoplasma genitalium; HSV-II: herpes simplex virus II.
Number of semen analyses performed and the incidence of infertility in different age groups. (a) Number of semen analyses. (b) Incidence of male infertility.