Gardnerella vaginalis in Symptomatic Men: Prevalence, Load, and Co-infections

TL;DR

Gardnerella vaginalis is commonly found in men with urethral symptoms and often occurs with other infections, suggesting it should be tested for in STI panels.

Contribution

This study is the first to quantify G. vaginalis prevalence and co-infection patterns in symptomatic men using PCR-based diagnostics.

Findings

Gardnerella vaginalis was detected in 21.5% of symptomatic men, more than any other pathogen.

81 patients had polymicrobial infections, often involving G. vaginalis with Ureaplasma species.

Over half of G. vaginalis cases had loads ≥10⁴ copies/mL, indicating clinical significance.

Abstract

Introduction

Gardnerella vaginalis, traditionally linked to bacterial vaginosis (BV) in women, is increasingly detected in men with urethral symptoms. Its clinical relevance and co-infection patterns remain poorly defined. This study assessed the prevalence, quantitative load, and co-infections of G. vaginalis, and other urogenital pathogens in symptomatic men.

Methods

We retrospectively analyzed 418 symptomatic male patients using multiplex polymerase chain reaction (PCR), targeting 21 pathogens. Pathogen loads were reported as qualitative or quantitative (<10⁴ or ≥10⁴ copies/mL).

Results

At least one pathogen was detected in 239 (57.2%) patients (95% CI: 52.4-61.9). G. vaginalis was most frequently identified: 90 (21.5%), followed by Chlamydia trachomatis: 63 (15.1%), Mycoplasma genitalium: 45 (10.8%), and Ureaplasma urealyticum: 38 (9.1%). Quantitative analysis showed 105 patients with G. vaginalis <10⁴ copies/mL and 90 with ≥10⁴ copies/mL. Polymicrobial infections occurred in 81 (19.5%) patients, most often involving G. vaginalis with U. urealyticum (n = 13) or U. parvum (n = 11).

Conclusions

G. vaginalis was the leading pathogen in symptomatic men, frequently present at clinically significant loads and in co-infections. These findings support its inclusion in routine male sexually transmitted infection (STI) diagnostic panels.

Tables

Table 1. Distribution of multiplex PCR results for each pathogenPCR, polymerase chain reaction

Pathogen	Negative, n(%)	Positive (qualitative), n(%)	Quantitative <10⁴, n(%)	Quantitative ≥10⁴, n(%)	Total positive, n(%)	95% CI
Candida spp.	446 (90.3%)	0 (0%)	46 (9.3%)	2 (0.4%)	48 (9.7%)	7.4-12.6
Chlamydia trachomatis	427 (86.3%)	68 (13.7%)	0 (0%)	0 (0%)	68 (13.7%)	11.0-17.1
Gardnerella vaginalis	299 (60.7%)	0 (0%)	105 (21.1%)	90 (18.2%)	195 (39.3%)	35.3-43.8
Herpes simplex virus (HSV) II	484 (97.8%)	11 (2.2%)	0 (0%)	0 (0%)	11 (2.2%)	1.2-3.9
Mycoplasma genitalium	446 (90.1%)	49 (9.9%)	0 (0%)	0 (0%)	49 (9.9%)	7.6-12.8
Mycoplasma hominis	468 (94.5%)	0 (0%)	11 (2.2%)	16 (3.3%)	27 (5.5%)	3.8-7.8
Neisseria gonorrhoeae	474 (95.8%)	21 (4.2%)	0 (0%)	0 (0%)	21 (4.2%)	2.8-6.4
Trichomonas vaginalis	493 (99.6%)	2 (0.4%)	0 (0%)	0 (0%)	2 (0.4%)	0.1-1.5
Ureaplasma urealyticum	429 (86.8%)	0 (0%)	27 (5.5%)	38 (7.7%)	65 (13.2%)	10.5-16.4

Table 2. Most common co-infection patterns among positive cases

Co-infection pattern	Number of patients, n(%)	95% CI
Gardnerella vaginalis + Ureaplasma urealyticum	13 (16.0%)	9.6-25.5
Chlamydia trachomatis + Mycoplasma genitalium	6 (7.4%)	3.4-15.2

Full text

Introduction

Sexually transmitted infections (STIs) remain a major global public health problem, with an estimated 374 million new cases of curable STIs occurring annually worldwide [1]. While Neisseria gonorrhoeae and Chlamydia trachomatis are the most frequently identified etiologic agents of male urethritis, advances in molecular diagnostics have revealed the role of non-classical and polymicrobial pathogens in urogenital infections [2,3].

Gardnerella vaginalis, a facultative anaerobic Gram-variable bacterium, is a well-established cause of bacterial vaginosis (BV) in women [4]. Historically regarded as a component of the normal vaginal microbiota, G. vaginalis has been increasingly detected in the male urogenital tract, particularly among men presenting with urethral symptoms [5]. Reported prevalence rates in male populations vary widely - from less than 1% to over 25% - depending on patient selection, sampling site, and diagnostic methodology [6,7].

The development of multiplex polymerase chain reaction (PCR) assays has improved the detection of G. vaginalis and other fastidious organisms, which are often missed by traditional culture or Gram staining [8,9]. These assays also allow for quantitative assessment of pathogen load, which may help distinguish between transient colonization and clinically significant infection.

Emerging evidence suggests that G. vaginalis may be involved in polymicrobial infections in men, frequently co-occurring with Ureaplasma spp. and Mycoplasma spp., potentially through synergistic interactions that enhance pathogenicity [10,11]. In addition, male carriage of G. vaginalis has been implicated in the recurrence of BV among female partners, underscoring its potential role in sexual transmission cycles [12,13].

Despite increasing recognition, the clinical significance of G. vaginalis in men remains poorly defined, and data on its prevalence, quantitative load, and co-infection patterns in symptomatic male populations are limited.

The aim of this study was to determine the prevalence and quantitative load of G. vaginalis and other urogenital pathogens in symptomatic men using multiplex PCR and to evaluate the frequency and patterns of co-infections in this population.

Materials and methods

Study design and population

This retrospective, descriptive study was conducted at the urology outpatient clinic of our institution between September 2022 and April 2025. The ethics committee approval for the study was obtained from the Memorial Hospital Local Ethics Committee on August 13, 2025, under number 2025/19. A total of 418 male patients presenting with symptoms suggestive of STIs were included. Only the first test result per patient was analyzed to avoid duplication.

Inclusion and exclusion criteria

Inclusion criteria were male patients aged ≥18 years and the presence of at least one of the following urogenital symptoms: urethral discharge, dysuria, penile pruritus, or penile discomfort. Exclusion criteria were asymptomatic individuals screened for other purposes; patients with incomplete clinical or laboratory records; duplicate samples from the same patient; patients who had used systemic or topical antibiotics, antifungals, or antiseptics within the last four weeks; patients with recent urethral catheterization, cystoscopy, or any other urological procedure within the previous three months; and detection of commensal flora organisms such as Lactobacillus spp., Staphylococcus spp., and Enterobacteriaceae spp., which were excluded from analysis.

Sample collection

Urethral swab samples were collected by trained healthcare professionals using sterile cotton-tipped applicators. The swab was inserted approximately 2-4 cm into the urethral meatus, rotated gently for several seconds, and then withdrawn. Samples were immediately placed into transport medium and sent to the microbiology laboratory for analysis. All samples were evaluated in the same laboratory in our hospital. In all patients, urethral Gram-stained smear microscopy was performed prior to PCR testing as part of routine clinical evaluation, although these results were not included in the current analysis. All samples were analyzed using a commercial multiplex real-time PCR system (Anyplex™ II STI-21; Seegene Inc., Seoul, Korea), performed on the CFX96™ Real-Time PCR Detection System (Bio-Rad Laboratories, Hercules, CA, USA).

Multiplex PCR analysis

All samples were tested using a commercial multiplex PCR assay capable of detecting 21 pathogens. The panel included G. vaginalis, C. trachomatis, N. gonorrhoeae, Ureaplasma urealyticum, Mycoplasma genitalium, Mycoplasma hominis, Trichomonas vaginalis, Candida spp., and Herpes simplex virus types 1 and 2 (HSV-1 and HSV-2).

Pathogen load was reported as negative, positive (qualitative only), quantitative <10⁴ copies/mL, or quantitative ≥10⁴ copies/mL. Commensal flora organisms were excluded from the final statistical analysis.

Statistical analysis

Data were entered into Microsoft Excel (Microsoft® Corp., Redmond, WA, USA) and analyzed using IBM SPSS Statistics for Windows, Version 21 (Released 2012; IBM Corp., Armonk, NY, USA). Categorical variables were expressed as frequencies and percentages, with 95% confidence intervals (CIs).

Results

Overall pathogen detection

A total of 418 symptomatic male patients were tested using multiplex PCR. At least one urogenital pathogen was detected in 239 patients, corresponding to an overall positivity rate of 57.2% (95% CI: 52.4-61.9). The remaining 179 (42.8%) patients had no detectable pathogens above the reporting threshold.

Pathogen distribution and quantitative loads

The most frequently identified organism was G. vaginalis, detected in 90 (21.5%) patients, followed by C. trachomatis in 63 (15.1%) patients, M. genitalium in 45 (10.8%) patients, and U. urealyticum in 38 (9.1%) patients. Less common pathogens included N. gonorrhoeae (n = 19; 4.5%), M. hominis (n = 16; 3.8%), HSV type II (n = 11; 2.6%), Candida spp. (n = 2; 0.5%), and T. vaginalis (n = 2; 0.5%).

Quantitative analysis revealed that G. vaginalis DNA was detected at a load of <10⁴ copies/mL in 105 patients, and ≥10⁴ copies/mL in 90 patients. For U. urealyticum, 27 patients had <10⁴ copies/mL, and 38 patients had ≥10⁴ copies/mL, while for U. parvum, 25 and 31 patients were in these respective categories. Detailed pathogen-specific quantitative data are also summarized in Table 1, categorized as negative, qualitative positive, quantitative <10⁴, or quantitative ≥10⁴.

Single and multiple infections

Of the 239 patients with at least one pathogen detected, 134 (56.1%) patients had single infections, while 105 (43.9%) patients had multiple pathogens detected. Overall, polymicrobial infections were identified in 81 patients (19.5% of the total cohort, 33.9% of positive cases). The most common co-infection patterns were G. vaginalis + U. urealyticum (n = 13, 16.0% of co-infected cases) and C. trachomatis + M. genitalium (n = 6, 7.4%). Table 2 lists the three most frequent co-infection patterns.

Discussion

In this large cohort of symptomatic men evaluated by multiplex PCR, G. vaginalis emerged as the most frequently detected urogenital pathogen, with a prevalence of 21.5% [2,6]. This detection rate is higher than those reported in several earlier studies, where prevalence in male urethral samples ranged from 4% to 15% [2,6]. Khosropour et al. [2] reported an 11% prevalence in symptomatic men, while Swidsinski et al. [5] observed an increase in G. vaginalis detection from 5% with culture to 18% with PCR, underscoring the improved sensitivity of molecular methods. The use of urethral swab sampling in our study, coupled with a broad multiplex PCR panel, likely contributed to the higher detection rate observed.

Notably, G. vaginalis was present as the sole pathogen in 7.4% of all cases and in combination with at least one other pathogen in 60% of G. vaginalis-positive cases. The most frequent co-infecting organisms were U. urealyticum and U. parvum, consistent with prior findings suggesting potential microbial synergy between G. vaginalis and other fastidious organisms [10,11]. Sarier et al. [4] demonstrated that G. vaginalis and M. hominis display synergistic pathogenicity in BV, which may have parallels in male urogenital infections.

The quantitative load analysis revealed that a substantial proportion of G. vaginalis and U. urealyticum detections were at high organism counts (≥10⁴ copies/mL). High bacterial loads have been associated with symptomatic disease and an increased likelihood of transmission [12,13]. The 2019 randomized controlled trial by Vodstrcil et al., published in the New England Journal of Medicine, demonstrated that treatment of male partners significantly reduced recurrence rates of BV in women, highlighting the potential role of men as reservoirs for reinfection [3]. In our cohort, G. vaginalis detections with bacterial loads below 10⁴ copies/mL were considered low-level colonization rather than definitive infection, consistent with previous reports emphasizing that clinically significant urethral infection is more likely when the microbial load exceeds this threshold.

While G. vaginalis is frequently detected, its pathogenic role in men is not fully defined. Some studies suggest transient colonization without overt symptoms [14,15], whereas case reports document its association with urethritis and prostatitis, particularly in immunocompromised patients [6,7]. The high prevalence and significant co-infection rate observed in our cohort support considering G. vaginalis as a relevant pathogen in symptomatic men, especially when detected at high quantitative loads.

Empirical treatment regimens for G. vaginalis typically include metronidazole or clindamycin [16,17]. However, reports of metronidazole resistance in male isolates raise concerns about the efficacy of standard protocols [5,18]. Where feasible, targeted therapy guided by susceptibility testing may improve treatment outcomes, particularly in recurrent or treatment-resistant cases [19].

Limitations

This study has several limitations. Its retrospective design precluded correlation of microbiological findings with detailed clinical outcomes or partner infection status. Antimicrobial susceptibility testing was not performed, and follow-up data on treatment response were unavailable. Additionally, the study population was limited to symptomatic men presenting to a single tertiary center, which may limit generalizability. The study included only symptomatic men; therefore, asymptomatic carriers and the potential reservoir role of G. vaginalis in the general male population were not evaluated. The lack of antimicrobial susceptibility testing and follow-up data limited our ability to assess response to treatment or recurrence. Another limitation is that cases with G. vaginalis loads <10⁴ copies/mL may represent colonization rather than true infection, which could have led to overestimation of its prevalence in symptomatic men.

Implications for practice and research

Our findings reinforce the need to expand diagnostic panels for male STIs to include G. vaginalis, particularly in cases of persistent or recurrent urethritis with negative results for classical pathogens. The frequent detection of high bacterial loads and co-infections supports the potential value of combination antimicrobial therapy. Future prospective studies should explore the longitudinal persistence of G. vaginalis in men, its role in male-to-female transmission, and optimal management strategies. Routine inclusion of G. vaginalis in diagnostic panels may be particularly useful for patients with persistent urethritis unresponsive to empirical therapy.

Conclusions

In this large cohort of symptomatic men, G. vaginalis was the most frequently detected urogenital pathogen, often present at high quantitative loads and in association with other sexually transmitted organisms. These findings suggest that G. vaginalis should be considered a relevant pathogen in male STI diagnostics, particularly when detected in high concentrations or in combination with other fastidious organisms.

Routine inclusion of G. vaginalis in multiplex PCR panels for men with urethral symptoms may improve diagnostic yield and guide more effective treatment strategies, including combination therapy in cases of polymicrobial infection. Further prospective studies are warranted to clarify its pathogenic role, persistence in the male urogenital tract, antimicrobial susceptibility patterns, and potential contribution to BV recurrence in female partners.