Influence of several uropathogenic microorganisms on human sperm motility parameters in vitro

Ji-Hong LIU, Hao-Yong LI, Zheng-Guo CAO, Yong-Fang DUAN, Yang LI, Zhang-Qun YE

Department of Urology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China

Asian J Androl 2002 Sep; 4: 179-182           

Keywords: Neisseria gonorrhoeae; Staphylococcus aureus; Staphylococcus epidermidis; Mycobacterium tuberculosis; sperm motility; computer-assisted sperm analysis

Abstract

Aim: The effects of certain uropathogenic microorganisms (Neisseria gonorrhoeae, Staphylococcus aureus, Staphylococcus epidermidis and Mycobacterium tuberculosis) on human sperm motility characteristics were studied in vitro. Methods: In 10 healthy fertile men, ejaculates were aseptically obtained by masturbation and with a swim-up technique, a sperm suspension of high motility and purity was obtained. Several uropathogenic bacteria were obtained from outpatients with genitourinary tract infections. The sperm suspension was incubated with the pathogens at a bacteria: sperm ratio of 50:1 at 37. The sperm mobility parameters were estimated with a computer-assisted sperm analyzer (CASA) provided with a multiple-exposure photography system (Madi Corp., Zhejiang, China). Measurements were carried out at 0, 2 and 4 hours of incubation. Results: Staphylococcus aureus significantly decreased the sperm motility and viability, but Staphylococcus epidermidis, Mycobacterium tuberculosis and Neisseria gonorrhoeae did not. Conclusion: Staphylococcus aureus has an inhibitory effect on human sperm motility in vitro.

Infection is one of the important causes of male infertility, which is usually curable [1-2]. The most widely studied genital microorganism in regard to male infertility was Escherichia coli of the O6 antigen serotype [3], which was also the principal microorganism causing prostatitis and epididymitis [4]. E. coli in vitro inhibited sperm motility and readily adhered to and agglutinated the sperm [5-6]. However, little is known about the influence of other uropathogenic bacteria on sperm motility. The present study was designed to observe the effect of Neisseria gonorrhoeae, Staphylococcus aureus, Staphylococcus epidermidis and Mycobacterium tuberculosis on human sperm motility by means of a computer-assisted sperm analyzer (CASA), based on the principle of multiple-exposure photography introduced by Makler et al [7].

2 Materials and methods

2.1 Sperm suspension

Ejaculates were obtained by masturbation into sterile plastic containers from healthy donors attending our artificial insemination donors programme after 4 days of sexual abstinence. They were free from genital tract infections and did not receive antibiotic therapy. Only ejaculates showing normal semen parameters according to the World Health Organization criteria [8] were used. For each experiment semen samples from 10 different donors were used.

To standardize the test and to establish an exact spermatozoa/microorganism ratio, after complete liquefaction and swim-up [9] treatment, the sperm suspension was diluted and adjusted to a concentration of 4010⁶ sperm/ml, a progressive motility (a+b) >40 %, viability >80 % and normal morphology >14 % [10] with Ham's F-10 medium (Sigma, USA).

2.2 Bacterial suspension

Four different uropathogenic microorganisms, Neisseria gonorrhoeae, Staphylococcus aureus, Staphylococcus epidermidis and Mycobacterium tuberculosis, were isolated and purified from the outpatients suffering from genital infections by standard culture technique. The growing colonies of bacteria from the culture were transferred into sterile tubes filled with Hams F-10 medium pre-heated to 37 to prepare living bacterial suspension at a concentration of 2.010⁹ colony forming units (cfu)/mL.

2.3 Incubation

Five aliquots (100 ml each) of the sperm suspension were prepared. One aliquot was added an equal amount of the Ham's F-10 medium and served as the control. Each of the other four aliquots was added an equal amount of one of the 4 microorganisms to be examined. The mixed suspensions (microorganism:sperm ratio 50:1) were then incubated at 37 for 4 hours in air.

2.4 Sperm motility assessment

The sperm motility parameters were analyzed by CASA at 0, 2 and 4 hours of incubation. The parameters included viability, percentage progressive motility, straight line velocity (VSL), curvilinear velocity (VCL) and average path velocity (VAP). Morphological observation was undertaken with a light microscope. Video recordings were made from eight different fields using a ?0 objective. At least 150 spermatozoa per sample were assessed for each group. The observation was made at room temperature.

2.5 Statistical analysis

Data were expressed in meanSEM. Statistical analyses were performed using the analysis of variance and the DUNNETT test for pairs by means of SAS software to determine the significance of differences between groups. P<0.05 was set as significant.

3 Results

All the microorganisms examined did not adhere to the spermatozoa and did not induce morphological changes or sperm agglutination.

3.2 Motility observation

Table 1 listed the effect of the microorganisms on the sperm motility. It could be seen that incubation with Staphylococcus aureus significantly decreased the progressive motility and other motility parameters (VSL, VCL, VAP, etc, P<0.05), particularly after 4 hours of incubation. It is interesting to note that the VCL dropped to less than 20 mm. It is well known that VCL >25 mm was used as the threshold value in predicting male fertility potential [11]. All other microorganisms did not significantly affect sperm motility parameters.

Table 1. Effect of different microorganisms on sperm motility and viability.

Genital tract infections affect spermatogenesis and sperm function in various ways, but their importance in male infertility is a matter of debate [12]. In recent years, it was indicated that pathogenic microorganisms could damage the sperm and cause infertility [13]. Diemer et al [6] demonstrated that E. coli (serotype O₆) inhibited sperm motility in vitro with the occurrence of sperm agglutination only at a microorganism/sperm ratio of more than 1; Electron microscopy revealed multiple adhesion of E. coli to spermatozoa, causing various ultrastructural damages. Many investigations indicated that genital tract infection with E. coli might be an important factor in male and female infertility [5].

Huwe et al [14] studied the influence of different uropathogenic microorganisms on human sperm motility parameters by means of CASA. They showed that E. coli, Pseudomonas aeruginosa and Candida albicans inhibited the sperm motility. Yin et al [15] showed that Streptococcus viridans decreased the progressive motility of sperm in vitro and at high concentrations the microorganism could induce sperm agglutination.

In accordance with our finding of the inhibitory effect of Staphylococcus aureus on progressive sperm motility, Jiang et al [16] found that the microorganism was the dominant flora in infertile men with a significant decrease in sperm motility. Staphylococcus aureus may produce various toxins and enzymes that may exert damaging effects on human sperm, but its mechanism of action needs further investigation.

Osegbe et al [17] demonstrated that epididymo-orchitis caused by Neisseria gonorrhoeae may result in azoospermia or oligospermia and infertility. Gomez et al [18] reported Neisseria gonorrhoeae could bind to human sperm, thus affecting the sperm function. However, the present study indicated that Neisseria gonorrhoeae had no significant impact on human sperm motility. Similarly, Swenson et al [19] did not see significant difference in sperm motility between semen sample infected by Neisseria gonorrhoeae and normal sample.

Recent studies showed that Staphylococcus epider-midis was the main cause of urethritis of the youth [20]. Zheng et al [21] demonstrated that 41.2% of chronic prostatitis were caused by Staphylococcus epidermidis and the sperm motility of these patients was significantly lower than that of healthy men. However, in our experi-ments, Staphylococcus epidermidis did not significantly affect the sperm motility.

Throughout the world, twenty per cent of patients with tuberculosis will develop an extrapulmonary manifestation over time, the most common site being the genitourinary tracts [22], leading to infertility. However so far, there is no information on the direct effect of tuberculous bacilli on human sperm. In the present study, Mycobacterium tuberculosis was found not to significantly affect sperm motility.

The in vitro results may only partially reflect the situations in vivo, therefore further investigations are needed to clarify the effect of the microorganisms on sperm motility and its mechanism of action.

References

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Correspondence to: Ji-Hong Liu, Ph.D., Department of Urology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.

Tel: +86-27-8366 2278, Fax: +86-27-8366 2591

E-mail: jhliu888@hotmail.com

Received 2002-04-02 Accepted 2002-04-18