Distribution and resistance patterns of male urethritis pathogens from a private South African laboratory

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The study also compared diagnostic performance between NAAT and culture and evaluated the relevance of syndromic management. This is the first large-scale South African private sector analysis of MUS pathogen profiles and AMR using NAAT and culture, providing critical evidence for diagnostic and management policy updates. Methods: A retrospective cross-sectional study was performed using laboratory data from 2012 to 2023, comprising over 61,000 samples. Organism prevalence and AMR rates were assessed. Diagnostic sensitivity and specificity of NAAT versus culture for Neisseria gonorrhoeae ( NG ) were calculated in matched samples. Results: NG and Chlamydia trachomatis were the most prevalent pathogens. NAAT demonstrated superior sensitivity, while culture yielded essential AMR data. NG showed elevated resistance patterns, but susceptibility to first-line antimicrobials remained high. Culture sensitivity and specificity relative to NAAT were 56% and 98.6%, respectively. Ureaplasma urealyticum showed elevated resistance versus global averages. The first cases of extensively drug-resistant (XDR) NG in South Africa were identified. Conclusion: NAAT improves MUS detection, while culture remains vital for AMR surveillance. Syndromic management remains effective, but rising resistance and XDR NG emergence warrant urgent attention. Urethritis Sexually transmitted infection Incidence Resistance Chlamydia trachomatis Neisseria Gonorrhoea Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 INTRODUCTION Male urethritis is most commonly contracted through infection during sexual intercourse and can be sub-classified into gonococcal (GU) and non-gonococcal urethritis (NGU) [ 1 – 3 ]. GU is caused by Neisseria gonorrhoeae (NG), the second most common bacterial sexually transmitted infection (STI) worldwide and the leading cause of male urethritis syndrome (MUS) in South Africa (70–80%) [ 4 , 5 ]. NGU is most commonly caused by Chlamydia trachomatis, Mycoplasma genitalium, Ureaplasma urealyticum and Trichomonas vaginalis [ 1 , 2 ]. These organisms penetrate the urethral epithelium, causing a pyogenic infection i.e. urethral discharge, the hallmark feature of urethritis. Nucleic acid amplification testing (NAAT) is considered the gold standard for aetiological testing in MUS and can be done on either first catch urine, or urethral swab specimens [ 6 ]. Microscopy culture and sensitivity (MC&S) testing can only be done on swabs [ 1 , 7 ]. Antimicrobial resistance patterns and spread pose a major risk to public health worldwide [ 5 ]. Resistance can develop quickly, as evidenced by a 0–22% increase in quinolone resistant NG over the course of 2023 in South Africa in [ 5 ]. Fortunately, Extensively Drug-Resistance (XDR) NG has never been identified in South Africa [ 5 ]. Impact on Public Health : Over 1 million new STI cases are diagnosed daily worldwide [ 8 ]. NG and Chlamydia Trachomatis are the most common causes of MUS in South Africa, with 4.5 million and 6 million new cases diagnosed in 2017 respectively [ 9 , 10 ]. Sub-Saharan Africa also has the highest incidence of NG in the world [ 11 ]. Despite this, incidence and aetiological data of MUS are severely lacking [ 11 ]. Antimicrobial resistance is also increasing rapidly, resulting in significant treatment challenges and global concern [ 2 , 5 ]. Relevant studies : There is minimal data on NAAT on urethral swabs for MUS in South Africa [ 10 ]. Existing original studies use small, specific populations in localised areas or conversely very large global or continental populations, limiting application in the South African context [ 6 , 7 , 8 , 10 – 13 ]. A study published in 2010 examined 300 men with MUS in a single family practice [ 10 ]. Their urine was subjected to NAAT and urethral swabs to gram staining. NAAT was only done on urine specimens, and the study population was small and from a concentrated demographic [ 10 ]. The 2018 GERMS SA surveillance review studied the aetiology of syndromic STI’s at three sentinel sites in South Africa. A total of only 258 MUS cases were included with no significant aetiological differences noted [ 16 ]. Other South African studies evaluated STI incidence in asymptomatic HIV positive populations attending primary health care clinics in a single province or ‘men who have sex with men’ (MSM) populations in South Africa using only first-pass urine specimens [ 17 , 18 ]. International studies include a Turkish study with a small population of only 91 MUS patients. Results, unfortunately, did not draw comparisons between urethral swabs and urine specimens, the only comparison done was between NAAT and culture results [ 8 , 12 ]. There are significant differences between sensitivity, specificity, and clinical application of first-pass urine and urethral swab specimens. The same is true for MC&S and NAAT. NAAT of urethral swabs has repeatedly proven to be the most sensitive means of testing for STI [ 19 – 21 ]. This has made NAAT the gold standard for STI diagnosis, with culture used for antimicrobial susceptibility testing [ 2 , 22 ]. Current Guidelines on MUS Management and Testing : In South Africa, STIs are primarily managed syndromically at the primary care level, targeting NG and C. trachomatis without requiring diagnostics [ 10 , 25 ]. Other relevant MUS pathogens include Mycoplasma genitalium and Trichomonas vaginalis, the latter needing alternative treatment [ 10 ]. Pathogen profiles may differ between public and private sectors, possibly necessitating adapted protocols [ 10 ]. Current guidelines recommend stat ceftriaxone 250mg IM and azithromycin 1g orally [ 2 , 23 ]. Persistent symptoms after seven days suggest ceftriaxone failure and require referral. The EAU supports doxycycline 100mg BD for 7 days as more effective than azithromycin, though adherence is lower [ 1 ]. MC&S is advised in treatment failure not due to reinfection or non-compliance [ 4 ]. Pettifor et al. (2000) supported syndromic management but highlighted the need for accessible diagnostics in low-resource settings [ 11 , 14 ]. STUDY AIM : Determine the prevalence of different causative organisms of MUS in South Africa. STUDY OBJECTIVES : Primary objective : To determine the prevalence of different causative organisms found on male urethral swabs in patients with urethritis in South Africa. Secondary objectives : To determine the rates of antimicrobial resistance in pathogens cultured on urethral swabs of male patients with urethritis in South Africa. To compare, within patients, pathogen identification using bacterial cultures to NAAT. To determine the relevance of current existing guidelines on syndromic management of STI’s and whether these are appropriate when compared to our findings. RESEARCH METHODS AND DESIGN Study Design : This was a retrospective cross-sectional study analysing data from 28 July 2012 to 31 December 2023 to describe the microbial aetiology of male urethritis and evaluate antimicrobial resistance patterns. Study Setting : Data were sourced from the central database of Lancet Laboratories, a private diagnostic laboratory network in South Africa. Study population and sampling strategy : Inclusion criteria: Males aged 16 years or older with MUS who submitted urethral swab or first catch urine specimens for NAAT and/or MC&S testing. Samples were collected and submitted by healthcare workers at various levels of care. All specimens were processed according to WHO-recommended protocols for STI diagnosis using molecular methods [ 6 , 24 ] and Clinical and Laboratory Standards Institute protocols [ 25 ]. Lancet laboratories were kind enough to supply anonymised categorised data for the above inclusion criteria in the form of a spreadsheet. Data Analysis: Five datasets were derived after cleaning and verification: Dataset 1: Culture-only results (n = 5 021) Dataset 2: NAAT panel (six organisms) (n = 1 055) Dataset 3: NAAT for N. gonorrhoeae only (n = 27 336) Dataset 4: NAAT for C. trachomatis only (n = 27 368) Dataset 5: Paired NAAT and culture data for N. gonorrhoeae (n = 191) Data were cleaned to remove duplicates and incomplete entries. All analyses were conducted in R version 4.4.2. Data were summarised using: Categorical variables (e.g., organism, resistance status): Frequency and percentage Continuous variables (e.g., age): Median, interquartile range (IQR), minimum and maximum To compare NAAT and culture detection of NG : Sensitivity, specificity, and accuracy of culture were calculated using NAAT as the reference standard Cohen’s Kappa Coefficient was used to assess method agreement [ 26 ]. Comparative resistance data between provinces and organism groups were analysed using Chi-squared tests. Ethical Considerations : The study was approved by the Human Research Ethics Committee of the University of the Witwatersrand (medical) (Appendix 1), with data access granted by the Lancet Laboratories Publication Committee (Appendix 2). No patient identifiers were provided, with all data being anonymised at source. Clearance certificate number: M240439 RESULTS Description of datasets After cleaning, four sets of data were used in the analysis: Dataset 1 (microbiology only) (primary objective & secondary objective 1): 5 021 individuals with urethral/penile/meatal swab cultures from 28 July 2012 to 29 Dec 2022. The age range was 16 to 97 years with median age being 37 years. 78.4% (n = 3 934) of samples were from Gauteng, and 15.5% from KZN (n = 781). Remaining samples were from: Free State: n = 52, Mpumalanga: n = 151, North West: n = 6, Northern Cape: n = 32, and Western Cape: n = 65. Dataset 2 (NAAT: Six bacterial species) (primary objective): 1 055 individuals with complete NAAT data for NG, Chlamydia trachomatis, Mycoplasma genitalium, Mycoplasma hominis, Trichomonas vaginalis, Ureaplasma parvum from urine (n = 953) or swab samples (n = 102) from 06 May 2015 to 28 Dec 2022. The age range was 16 to 84 years with a median of 35 years. Ninety seven percent (n = 1 027) of the samples were from Gauteng. Remaining samples were from: KZN = 7, Mpumalanga: n = 3, North West: n = 1, Northern Cape: n = 1, and Western Cape: n = 16. Dataset 3 (NAAT : NG only) (primary objective): 27 366 individuals with NAAT data for NG DNA from urine samples (n = 26 982) or swab samples (n = 384) from 17 Dec 2014 to 31 Dec 2022 (8 years). The age range was 16 to 90 years with a median of 35. Eighty five percent (n = 23 210) of samples were from Gauteng, 8%, were from KZN (n = 2 281), and remaining samples from: Eastern Cape: n = 1, Free State: n = 288, Mpumalanga: n = 585, North West: n = 260, Northern Cape: n = 103, and Western Cape: n = 638. Dataset 4 (NAAT: Assessed for Chlamydia trachomatis only) (primary objective) 27 368 individuals with NAAT data for Chlamydia trachomatis from urine samples (n = 26 984) or swab samples (n = 384) from 17 Dec 2014 to 31 Dec 2022 (8 years). The age range was 6 to 90 years with a median of 35. Eighty five percent (n = 23 211) were from Gauteng, 8.3% were from KZN (n = 2 281) with remaining samples from: Eastern Cape: n = 1, Free State: n = 288, Mpumalanga: n = 585, North West: n = 260, Northern Cape: n = 103, and Western Cape: n = 639. Dataset 5 (combined culture and NAAT data) (secondary objective 2): 191 individuals with both NAAT and culture data for NG from 27 September 2013 to 12 December 2022 (~ 9 years). The age range was 17 to 77 years with a median of 34. Eighty six percent (n = 165) came from Gauteng with remaining samples from: KZN: n = 20, Mpumalanga: n = 2, and Western Cape: n = 4. ~35% of NAAT assays detected NG , and ~ 19% of cultures detected NG . Primary objective : To determine the prevalence of different causative organisms in MUS in South Africa using bacterial culture and NAAT. Dataset 1: 5,021 cultures (2012–2022); 58% showed no growth. NG was most frequent (12%), followed by Candida and E. coli. In 5,021 cultures, 58% were negative; 42% yielded organisms. NG was most common (12%). Neisseria gonorrhoeae (NG) was the most frequently isolated organism (~ 12%), occurring 3.4 times more often than the next most common, Candida. Escherichia coli was the most common bacterial species after NG (3.3%). Ureaplasma urealyticum, a recognised STI pathogen, was the sixth most common isolate (2%). Dataset 2: 1,055 NAATs for six species (2015–2022); 80% were negative. CT, UP, and MG were most common positives. Of 1,055 NAAT samples, NG was 2.1%. Most positives were non-gonococcal. Dataset 3: 27,336 NAATs for NG (2014–2022); 3.8% positive. Among 27,336 NAATs for NG, 3.8% were positive. Dataset 4: 27,368 NAATs for CT (2014–2022); 7.1% positive. Of 27,368 tested for CT, 7.1% were positive. Secondary objective 1 : Determine the frequency of antimicrobial resistance in STI pathogens cultured from urethral swabs of male patients with urethritis in South Africa. Determine the frequency of antimicrobial resistance in STI pathogens cultured from urethral swabs of male patients with urethritis in South Africa. Neisseria gonorrhoeae showed high sensitivity to azithromycin (89.3%) and cefixime (93.8%), but markedly lower rates for ciprofloxacin (34.9%), penicillin (23.9%), and tetracycline (13.1%) (Fig. 3 ). Ureaplasma urealyticum demonstrated moderate sensitivity to erythromycin (75.5%) and tetracycline (61.2%) (Fig. 3 ). Provincial variation in NG sensitivity (Fig. 4 ): Antibiotic sensitivity data were analysed for Gauteng, KZN, and Western Cape (≥ 10 NG cultures per antibiotic). Azithromycin data were only available from Gauteng; KZN lacked tetracycline data. Cefixime sensitivity was high in Gauteng (94%) and KZN (95.2%), but lower in WC (78.6%). Ciprofloxacin susceptibility was poor overall but highest in KZN (51.2%) compared to Gauteng (27.9%) and WC (38.5%). Penicillin susceptibility followed a similar trend: KZN (29.5%) > Gauteng (17.6%) > WC (14.2%). Tetracycline resistance was high in both Gauteng (88.1%) and WC (54.5%). Limitations and combination therapy analysis (Fig. 5 ): Only Gauteng had sufficient Ureaplasma urealyticum data for analysis, limiting interprovincial comparison. Current first-line syndromic management includes ceftriaxone, azithromycin, and doxycycline. Doxycycline was not tested against NG and was used in only two U. urealyticum cultures—each as the sole antibiotic tested. No NG cultures included ceftriaxone, but azithromycin and cefixime were assessed. Among NG isolates with both results, 84% were sensitive to both drugs, 9.7% were cefixime-sensitive but azithromycin-resistant, 5.2% showed the reverse, and dual resistance was rare (1.1%). Only Gauteng and WC contributed to this analysis, with just four cases from WC, precluding meaningful provincial comparison. Secondary objective 2: Culture vs NAAT for NG detection ( Fig. 6 ) : A total of 191 patients had matched NAAT and culture data for Neisseria gonorrhoeae. Using NAAT as the reference standard, culture showed high specificity but limited sensitivity. Culture accuracy was 87.4% (95% CI: 81.9–91.8), sensitivity 56%, and specificity 98.6%. Cohen’s Kappa was 0.63, indicating moderate-to-good agreement between methods. Culture was negative in 84.3% (n = 161) and positive in 15.7% (n = 30). NAAT detected NG in 50 cases (26.2%), but 22 of these were missed by culture. Only two samples were culture-positive but NAAT-negative, highlighting a high false-negative rate for culture. DISCUSSION The study provides a detailed overview of aetiology and resistance patterns of sexually transmitted MUS in South Africa’s private healthcare sector over a ten-year period. The study incorporates both culture-based and molecular diagnostic techniques, resulting in one of the largest and most comprehensive datasets of its kind in South Africa. The findings have important implications for current syndromic management algorithms and diagnostic modalities. Pathogen Distribution : NG was the most commonly identified pathogen (~ 12%). Urine is less invasive and easier to handle than swabs, influencing clinician preference. These findings support molecular diagnostics for NGU screening. Antimicrobial Resistance Patterns : Neisseria gonorrhoeae The AMR profile of NG demonstrated marked resistance variability. Lowest resistance was observed for cefixime (5.8%) and azithromycin (10.7%), while ciprofloxacin (65.1%), penicillin (76.1%), and tetracycline (86.9%) showed unacceptably high resistance. Quinolone resistance rose sharply from 0–22% in KZN in 2023 to 65.1% [ 5 ], echoing global trends and reinforcing the exclusion of ciprofloxacin and tetracycline from empiric regimens [ 34 , 35 ]. These trends align with the WHO’s classification of NG AMR as a high-priority threat [ 36 ]. Regional differences were notable: Cefixime susceptibility in WC was 78.6%, compared to 94% in Gauteng and 95.2% in KZN, suggesting regional clonal variation or prescribing differences. Prior AMR data were limited to Gauteng, KZN, Eastern Cape, and WC [ 5 ], yet resistance appears to be worsening. Tetracycline-resistant NG (TRNG) is now globally prevalent [ 37 ]. In South Africa, TRNG increased from 73% in 2011 to 86.9% in the present data [ 37 ]. Prior studies noted a preference for oral cephalosporins (cefixime) in high-income settings versus intramuscular ceftriaxone in lower-income regions [ 37 ]. This distinction is evident locally: Cefixime use dominates in the private sector, likely due to ease of oral administration and less price sensitivity, whereas ceftriaxone remains standard in the public sector. Despite cefixime not being recommended in MUS guidelines or listed on the government’s essential medicines list [ 38 ], its use persists in the private sector, even with greater resistance than ceftriaxone [ 39 ]. Private providers are thus advised to align with syndromic protocols despite convenience or patient preference for oral therapy. Comparison to Previous Studies on Neisseria gonorrhoeae in South Africa The AMR patterns largely reflect, but also diverge from, those reported by Yakobi et al. (2024). As similarly observed, high resistance to ciprofloxacin, penicillin, and tetracycline was found across Gauteng, KZN, and WC. However, ciprofloxacin resistance was significantly higher in the current study (65.1%) compared to Yakobi et al.’s findings (23% in KZN, 14% in Gauteng, 100% in WC), possibly due to evolving resistance or private sector prescribing patterns [ 5 ]. Penicillin susceptibility was low (23.9%), aligning with resistance reported in Gauteng (23%) and KZN (25%) by Yakobi et al. [ 5 ]. In WC, susceptibility was even lower (14.2%), although Yakobi et al. did not provide comparative data for this province. Tetracycline resistance was extensive (86.9%), exceeding Yakobi et al.’s national rate (30%) (5). Gauteng showed 88.1% resistance, WC 54.5%, compared to Yakobi’s 61% in Gauteng and 36% in KZN [ 5 ]. In contrast, azithromycin (89.3%) and cefixime (93.8%) retained high susceptibility, consistent with Yakobi et al.’s low resistance rates (azithromycin: 17% in KZN, 1% in Gauteng; cefixime: ≤0.6%) [ 5 ]. However, cefixime susceptibility was lower in WC (78.6%) than in Gauteng and KZN (94–95%), warranting surveillance. Dual susceptibility to azithromycin and cefixime was found in 84% of isolates, supporting current treatment guidelines. Notably, this is the first report of XDR N. gonorrhoeae in South Africa [ 5 ], with a prevalence of 1.1%. XDR NG is defined as resistance to both recommended therapies (cephalosporin and azithromycin) plus resistance to two or more other antimicrobials (penicillin, tetracycline, erythromycin) [ 40 ]. Among the three XDR cases, one isolate was sensitive to penicillin, one to tetracycline, and one resistant to all five agents tested. These findings highlight provincial AMR differences, underscoring the need for regional surveillance. Ureaplasma urealyticum : Resistance in Ureaplasma urealyticum (UU) is notable due to its lack of a cell wall, rendering it naturally resistant to all cell-wall-targeting antibiotics, particularly β-lactams [ 41 ], which limits treatment options and exacerbates resistance concerns. Resistance to first-line macrolides and tetracyclines was prominent, especially tetracycline (38.8%), while susceptibility to erythromycin remained relatively preserved at 75.5% [ 41 ]. This resistance markedly exceeds global averages reported by Ramaloko et al. (2025), where macrolide and tetracycline resistance were 19% and 8%, respectively [ 41 ]. The observed tetracycline resistance is roughly fivefold higher than global estimates (38.8% vs 8%), raising concerns about empiric doxycycline use in syndromic STI management [ 41 , 42 ]. Similarly, the 24.5% erythromycin resistance rate surpasses the global mean of 19%, indicating worsening macrolide resistance in local Ureaplasma strains, potentially driven by widespread azithromycin use in syndromic protocols [ 38 , 41 ]. Historically, UU has been considered more macrolide-susceptible than Mycoplasma genitalium, but this study suggests this may no longer hold true in South African private healthcare settings. While fluoroquinolone resistance and multidrug resistance (MDR) were not assessed here, these remain critical areas for future surveillance. Given high resistance rates to two primary antimicrobial classes, MDR UU may be circulating undetected in South Africa. Antimicrobial resistance studies remain limited nationally [ 5 ]. Provinces such as Mpumalanga and North West have never undergone NG AMR surveillance, and many regions lack NG susceptibility data since the introduction of syndromic management in the 1990s [ 41 ]. Enhanced routine STI aetiological monitoring is urgently needed to update syndromic treatment regimens [ 5 ], despite challenges posed by budget and logistical constraints. This is vital for South Africa’s progress toward the WHO target of 90% STI reduction by 2030 [ 31 , 36 ]. Europe’s modest 3% annual decline in GU incidence since this target was set illustrates the difficulty of this task [ 28 ]. Comparison of Culture vs NAAT : In 191 samples tested for Neisseria gonorrhoeae (NG) by both NAAT and culture, culture demonstrated a sensitivity of 56% and specificity of 98.6%, with Cohen’s κ = 0.63, indicating good agreement. In contrast, Serra-Pladevall et al. (2015) reported higher culture sensitivity (86.2%) and specificity (99.8%) [ 43 ], their study treated urine specimens as equivalent to urethral swabs, differing from the present methodology [ 43 ]. NAAT in their study showed sensitivity and specificity of 98.7% and 100% respectively [ 43 ]. Thus, while specificities are comparable, the markedly lower culture sensitivity here suggests up to 44% of NG infections may be missed. These findings confirm culture’s known limitations as a low-sensitivity, high-specificity diagnostic tool for NG [ 42 ]. The moderate agreement and poor sensitivity highlight that culture alone is insufficient and may lead to under-treatment. Strengths and Limitations of Study : This is the first large-scale private sector analysis of MUS pathogens and resistance in South Africa. Limitations include reliance solely on laboratory results without access to specimen collection methods, patient history, or clinical symptoms. Standardisation across this large, geographically diverse cohort was not feasible. Additionally, the study lacked ceftriaxone resistance data, may be affected by selection bias, and did not include longitudinal follow-up or treatment outcomes. Conclusions The study supports ongoing dual therapy for MUS in South Africa but raises concerns about the long-term use of azithromycin due to emerging resistance. It also advises against routine doxycycline use in syndromic management owing to high resistance rates. The study highlights the limitations of culture in diagnosing NGU and underscores the increasing value of NAAT-based multiplex panels, especially in atypical or persistent cases. However, culture remains essential for antimicrobial resistance profiling. The unprecedented finding of XDR NG in South Africa is of great concern to public health. Declarations The authors have no financial or personal relationships that may have inappropriately influenced the content of this thesis. No conflicts of interest are present. Competing interests: The authors have no conflicts of interest to declare that are relevant to the content of this article. Funding: No funding was received for conducting this study. Disclaimer: The views expressed in this article are those of the author and not of any associated institution. Author contributions: Dr. Matthew Snyman is the corresponding author for this study, and conducted all communication with outside parties, conducted the literature review, collected and analysed data, drew conclusions and wrote the final paper as an original work. Dr. Marietha Nel and Dr. Winston Padayachee jointly supervised this study in equal contribution, providing valuable insight, advice and assistance in proofreading and correcting the final write-up. DEDICATION: To my family, for their unwavering support, to my mentors, who guided me and to my patients who have inspired me. ETHICS APPROVAL: University of the Witwatersrand Human Research Ethics Committee (Medical) approved this study and designated M240439 as a reference number ACKNOWLEDGEMENTS: Dr Marietha Nel and Dr Winston Padayachee, for your supervision and guidance overseeing my execution of this research and write-up. Prof Peter Kamerman for assistance with the statistics and data analysis of results and for your invaluable insights and guidance. Professor Ahmed Adam for your guidance and expert advice Lancet Laboratories for kindly providing access to their extensive database of results References EAU Guidelines Office EAU Guidelines. Edn. presented at the EAU Annual Congress Milan, Italy 2023. [Internet]. Arnhem, The Netherlands.: EAU Guidelines Office; 2023 [cited 2023 Sep 24]. Available from: https://uroweb.org/guidelines/urological-infections/chapter/citation-information The State of World Health (1995) : World Health Organization. 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Available from: https://www.eurosurveillance.org/content/ 10.2807/1560-7917.ES.2024.29.9.2300226 Michalow J, Hall L, Rowley J, Anderson RL, Hayre Q, Chico RM et al (2024) Prevalence of chlamydia, gonorrhoea, and trichomoniasis among male and female general populations in sub-Saharan Africa from 2000–2024: A systematic review and meta-regression analysis [Internet]. Infectious Diseases (except HIV/AIDS); [cited 2025 Apr 16]. Available from: http://medrxiv.org/lookup/doi/10.1101/2024.12.16.24319070 Unemo M, Ballard R, Ison C, Lewis D, Ndowa F, Peeling R, et al. Laboratory diagnosis of sexually transmitted infections, including human immunodeficiency virus / edited by Magnus Unemo … et al] [Internet]. Geneva: World Health Organization; 2013 [cited 2025 Apr 21]. Available from: https://iris.who.int/handle/10665/85343 Rowley J, Vander Hoorn S, Korenromp E, Low N, Unemo M, Abu-Raddad LJ et al (2019) Chlamydia, gonorrhoea, trichomoniasis and syphilis: global prevalence and incidence estimates, 2016. Bull World Health Organ 97(8):548–562P Kularatne R, Maseko V, Mahlangu P, Muller E, Kufa T (2022) Etiological Surveillance of Male Urethritis Syndrome in South Africa: 2019 to 2020. Sex Transm Dis 49(8):560–564 Van Der Pol B, Ferrero DV, Buck-Barrington L, Hook E, Lenderman C, Quinn T et al (2001) Multicenter Evaluation of the BDProbeTec ET System for Detection of Chlamydia trachomatis and Neisseria gonorrhoeae in Urine Specimens, Female Endocervical Swabs, and Male Urethral Swabs. J Clin Microbiol 39(3):1008–1016 Moodley P, Sturm AW (2000) Sexually transmitted infections, adverse pregnancy outcome and neonatal infection. Semin Neonatol 5(3):255–269 Manhart LE, Broad JM, Golden MR (2011) Mycoplasma genitalium: Should We Treat and How? Clin Infect Dis 53(suppl3):S129–S142 Durukan D, Read TRH, Murray G, Doyle M, Chow EPF, Vodstrcil LA et al (2020) Resistance-Guided Antimicrobial Therapy Using Doxycycline–Moxifloxacin and Doxycycline–2.5 g Azithromycin for the Treatment of Mycoplasma genitalium Infection: Efficacy and Tolerability. Clin Infect Dis 71(6):1461–1468 Global Health Sector Strategies on, Respectively HIV (2022) Viral Hepatitis and Sexually Transmitted Infections for the Period 2022–2030, 1st edn. World Health Organization, Geneva, p 1 Lewis DA (2011) Antimicrobial-resistant gonorrhoea in Africa: An important public health threat in need of a regional gonococcal antimicrobial surveillance programme. South Afr J Epidemiol Infect 26(4):215–220 Standard Treatment Guidelines and Essential Medicines List | Department of Health Knowledge Hub [Internet]. [cited 2025 Apr 21]. Available from: https://knowledgehub.health.gov.za/content/standard-treatment-guidelines-and-essential-medicines-list Le Van A, Rahman N, Sandy R, Dozier N, Smith HJ, Martin MJ et al Common Patterns and Unique Threats in Antimicrobial Resistance as Demonstrated by Global Gonococcal Surveillance. Emerg Infect Dis [Internet]. 2024 Oct [cited 2025 Apr 16];30(14). Available from: https://wwwnc.cdc.gov/eid/article/30/14/24-0296_article Martin I, Sawatzky P, Allen V, Lefebvre B, Hoang L, Naidu P et al (2019) Multidrug-resistant and extensively drug-resistant Neisseria gonorrhoeae in Canada, 2012–2016. Can Commun Dis Rep 45(2/3):45–53 Ramaloko WT, Maningi NE, Osei Sekyere J (2025) Global prevalence, resistance rates, and underlying resistance mechanisms of clinical Mycoplasma and Ureaplasma species. J Appl Microbiol 136(1):lxae308 Taylor-Robinson D, Jensen JS (2011) Mycoplasma genitalium: from Chrysalis to Multicolored Butterfly. Clin Microbiol Rev 24(3):498–514 Serra-Pladevall J, Caballero E, Roig G, Juvé R, Barbera MJ, Andreu A (2015) Comparison between conventional culture and NAATs for the microbiological diagnosis in gonococcal infection. Diagn Microbiol Infect Dis 83(4):341–343 Additional Declarations No competing interests reported. Supplementary Files Appendix.docx Cite Share Download PDF Status: Published Journal Publication published 11 Dec, 2025 Read the published version in World Journal of Urology → Version 1 posted Editorial decision: Revision requested 29 Oct, 2025 Reviews received at journal 24 Oct, 2025 Reviewers agreed at journal 06 Oct, 2025 Reviewers invited by journal 30 Jul, 2025 Editor assigned by journal 21 Jul, 2025 Submission checks completed at journal 21 Jul, 2025 First submitted to journal 09 Jul, 2025 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7085306","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":495015731,"identity":"e8bfdfa2-9c3e-48ad-ba99-03ee464a6fd1","order_by":0,"name":"Matthew Dylan Snyman","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA8klEQVRIiWNgGAWjYJACZgSzAsRlbiCkgbEZwTkDFiBFC2MbmMSvRbf9/PHHBRWH7c0lshM/F86rjeZvB2r5UbENpxazM8mMzTPOHE7cOSN3s/TMbcdzZxxmbGDsOXMbt5YDQC28bbcTDG7kbpDm3XYstwGohZmxDY+W84+BWv7dtgdq2fybd86x3PkEtdwA2dJwm3HDjdxt0rwNNbkbCGt5bDib59j/xA1n3m6z5jl2IHcjUMtBvH45n/jgM09Nmr3B8dzNt3lq6nLnnT988MGPCtxaEEAgAUQeBrMPEKEeCPjB6uqIUzwKRsEoGAUjCgAAjNNibijZbGIAAAAASUVORK5CYII=","orcid":"","institution":"University of the Witwatersrand","correspondingAuthor":true,"prefix":"","firstName":"Matthew","middleName":"Dylan","lastName":"Snyman","suffix":""},{"id":495015732,"identity":"82f07845-c3bf-4e94-92f8-11f53671c62c","order_by":1,"name":"Marietha Nel","email":"","orcid":"","institution":"University of the Witwatersrand","correspondingAuthor":false,"prefix":"","firstName":"Marietha","middleName":"","lastName":"Nel","suffix":""},{"id":495015733,"identity":"78382d6b-7695-4a7b-9b9a-88e084f359f9","order_by":2,"name":"Winston Padayachee","email":"","orcid":"","institution":"University of the Witwatersrand","correspondingAuthor":false,"prefix":"","firstName":"Winston","middleName":"","lastName":"Padayachee","suffix":""}],"badges":[],"createdAt":"2025-07-09 15:23:32","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7085306/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7085306/v1","draftVersion":[],"editorialEvents":[{"content":"https://doi.org/10.1007/s00345-025-06117-y","type":"published","date":"2025-12-11T15:57:00+00:00"}],"editorialNote":"","failedWorkflow":false,"files":[{"id":88226658,"identity":"cfc4ab37-aef2-429b-b499-26227cf6a743","added_by":"auto","created_at":"2025-08-04 08:41:33","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":175054,"visible":true,"origin":"","legend":"\u003cp\u003eIsolation rates of bacterial species (or Candida) from 2,398 cultured specimens. Only the 11 species with ≥1% frequency are shown. “STI” denotes sexually transmitted infection; “Gonococcal” refers to Neisseria gonorrhoeae\u003c/p\u003e","description":"","filename":"image1.png","url":"https://assets-eu.researchsquare.com/files/rs-7085306/v1/92ad5d4743b31e6a988d2a99.png"},{"id":88226656,"identity":"26003a8e-7414-4ed2-8862-eba088165582","added_by":"auto","created_at":"2025-08-04 08:41:33","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":107818,"visible":true,"origin":"","legend":"\u003cp\u003eDetection rates of bacterial genetic material by NAAT in 1,055 specimens. “Other bacteria” includes Mycoplasma genitalium, Mycoplasma hominis, Trichomonas vaginalis, and Ureaplasma parvum. “Chlamydia” refers to Chlamydia trachomatis; “Gonococcal” to Neisseria gonorrhoeae\u003c/p\u003e","description":"","filename":"image2.png","url":"https://assets-eu.researchsquare.com/files/rs-7085306/v1/5d0a12b05cdbed16a48fc2f9.png"},{"id":88226978,"identity":"dac76167-3c1f-4b13-8c36-89f926fbb275","added_by":"auto","created_at":"2025-08-04 08:49:33","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":122397,"visible":true,"origin":"","legend":"\u003cp\u003eAntibiotic resistance in Neisseria gonorrhoeae (top) and Ureaplasma urealyticum (bottom). Numbers in parentheses indicate test counts per antibiotic. Sensitivity categories: VS (very susceptible), S (susceptible), I (intermediate), R (resistant), MR (moderately resistant), VR (very resistant), TF (treatment failure)\u003c/p\u003e","description":"","filename":"image3.png","url":"https://assets-eu.researchsquare.com/files/rs-7085306/v1/6c34e8993c97fd936142b86d.png"},{"id":88226661,"identity":"3fc668e5-d3a4-41e1-9a94-0151c1af1c78","added_by":"auto","created_at":"2025-08-04 08:41:33","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":170244,"visible":true,"origin":"","legend":"\u003cp\u003eNeisseria gonorrhoeae antibiotic resistance by province. Numbers in parentheses indicate the number of tests per antibiotic. Sensitivity categories: VS (very susceptible), S (susceptible), I (intermediate), R (resistant), MR (moderately resistant), VR (very resistant), TF (treatment failure)\u003c/p\u003e","description":"","filename":"image4.png","url":"https://assets-eu.researchsquare.com/files/rs-7085306/v1/7407a4cd6312d978eb6a4b2d.png"},{"id":88226668,"identity":"4deadec2-0629-459a-a55d-84aa6747f0a8","added_by":"auto","created_at":"2025-08-04 08:41:33","extension":"png","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":94970,"visible":true,"origin":"","legend":"\u003cp\u003eNeisseria gonorrhoeae resistance patterns based on sensitivity to cefixime and azithromycin, tested on separate cultures from the same swab. Each bar represents a resistance/sensitivity combination (S = sensitive, R = resistant); numbers in parentheses indicate the frequency of each combination. Note: antibiotics were not tested simultaneously on the same culture\u003c/p\u003e","description":"","filename":"image5.png","url":"https://assets-eu.researchsquare.com/files/rs-7085306/v1/c5573c49be7da258f14bbc1b.png"},{"id":88226665,"identity":"57fe2ac0-2350-4bbe-b461-f90011355f65","added_by":"auto","created_at":"2025-08-04 08:41:33","extension":"png","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":92110,"visible":true,"origin":"","legend":"\u003cp\u003eMosaic plot of Neisseria gonorrhoeae detection by culture and NAAT (n = 191). Block areas reflect sample counts: top left (n = 22), top right (n = 139), bottom left (n = 28), and bottom right (n = 2). Colours represent culture outcomes (light = negative, dark = positive)\u003c/p\u003e","description":"","filename":"image6.png","url":"https://assets-eu.researchsquare.com/files/rs-7085306/v1/dd810aef8fd02666c44402b3.png"},{"id":98243493,"identity":"1284418d-37c4-4067-821a-eb25b81ce4c6","added_by":"auto","created_at":"2025-12-15 16:06:43","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":1435058,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7085306/v1/890c685a-2c62-4bd0-b5e8-1fe31edbdba9.pdf"},{"id":88227966,"identity":"9637d4f6-d868-4063-bace-c78366e6fc8a","added_by":"auto","created_at":"2025-08-04 08:57:33","extension":"docx","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":481518,"visible":true,"origin":"","legend":"","description":"","filename":"Appendix.docx","url":"https://assets-eu.researchsquare.com/files/rs-7085306/v1/338c63387e38517ee3108c36.docx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Distribution and resistance patterns of male urethritis pathogens from a private South African laboratory","fulltext":[{"header":"INTRODUCTION","content":"\u003cp\u003eMale urethritis is most commonly contracted through infection during sexual intercourse and can be sub-classified into gonococcal (GU) and non-gonococcal urethritis (NGU) [\u003cspan additionalcitationids=\"CR2\" citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. GU is caused by Neisseria gonorrhoeae (NG), the second most common bacterial sexually transmitted infection (STI) worldwide and the leading cause of male urethritis syndrome (MUS) in South Africa (70\u0026ndash;80%) [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. NGU is most commonly caused by Chlamydia trachomatis, Mycoplasma genitalium, Ureaplasma urealyticum and Trichomonas vaginalis [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. These organisms penetrate the urethral epithelium, causing a pyogenic infection i.e. urethral discharge, the hallmark feature of urethritis. Nucleic acid amplification testing (NAAT) is considered the gold standard for aetiological testing in MUS and can be done on either first catch urine, or urethral swab specimens [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. Microscopy culture and sensitivity (MC\u0026amp;S) testing can only be done on swabs [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eAntimicrobial resistance patterns and spread pose a major risk to public health worldwide [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. Resistance can develop quickly, as evidenced by a 0\u0026ndash;22% increase in quinolone resistant NG over the course of 2023 in South Africa in [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. Fortunately, Extensively Drug-Resistance (XDR) NG has never been identified in South Africa [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e].\u003c/p\u003e\u003cp\u003e\u003cb\u003eImpact on Public Health\u003c/b\u003e:\u003c/p\u003e\u003cp\u003eOver 1\u0026nbsp;million new STI cases are diagnosed daily worldwide [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. NG and Chlamydia Trachomatis are the most common causes of MUS in South Africa, with 4.5\u0026nbsp;million and 6\u0026nbsp;million new cases diagnosed in 2017 respectively [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e, \u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Sub-Saharan Africa also has the highest incidence of NG in the world [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. Despite this, incidence and aetiological data of MUS are severely lacking [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e]. Antimicrobial resistance is also increasing rapidly, resulting in significant treatment challenges and global concern [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e].\u003c/p\u003e\u003cp\u003e\u003cb\u003eRelevant studies\u003c/b\u003e:\u003c/p\u003e\u003cp\u003eThere is minimal data on NAAT on urethral swabs for MUS in South Africa [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Existing original studies use small, specific populations in localised areas or conversely very large global or continental populations, limiting application in the South African context [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e, \u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan additionalcitationids=\"CR11 CR12\" citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eA study published in 2010 examined 300 men with MUS in a single family practice [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Their urine was subjected to NAAT and urethral swabs to gram staining. NAAT was only done on urine specimens, and the study population was small and from a concentrated demographic [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. The 2018 GERMS SA surveillance review studied the aetiology of syndromic STI\u0026rsquo;s at three sentinel sites in South Africa. A total of only 258 MUS cases were included with no significant aetiological differences noted [\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. Other South African studies evaluated STI incidence in asymptomatic HIV positive populations attending primary health care clinics in a single province or \u0026lsquo;men who have sex with men\u0026rsquo; (MSM) populations in South Africa using only first-pass urine specimens [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e, \u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e]. International studies include a Turkish study with a small population of only 91 MUS patients. Results, unfortunately, did not draw comparisons between urethral swabs and urine specimens, the only comparison done was between NAAT and culture results [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e, \u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThere are significant differences between sensitivity, specificity, and clinical application of first-pass urine and urethral swab specimens. The same is true for MC\u0026amp;S and NAAT. NAAT of urethral swabs has repeatedly proven to be the most sensitive means of testing for STI [\u003cspan additionalcitationids=\"CR20\" citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e\u0026ndash;\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e]. This has made NAAT the gold standard for STI diagnosis, with culture used for antimicrobial susceptibility testing [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e].\u003c/p\u003e\u003cp\u003e\u003cb\u003eCurrent Guidelines on MUS Management and Testing\u003c/b\u003e:\u003c/p\u003e\u003cp\u003eIn South Africa, STIs are primarily managed syndromically at the primary care level, targeting NG and C. trachomatis without requiring diagnostics [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e, \u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. Other relevant MUS pathogens include Mycoplasma genitalium and Trichomonas vaginalis, the latter needing alternative treatment [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Pathogen profiles may differ between public and private sectors, possibly necessitating adapted protocols [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eCurrent guidelines recommend stat ceftriaxone 250mg IM and azithromycin 1g orally [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]. Persistent symptoms after seven days suggest ceftriaxone failure and require referral. The EAU supports doxycycline 100mg BD for 7 days as more effective than azithromycin, though adherence is lower [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e]. MC\u0026amp;S is advised in treatment failure not due to reinfection or non-compliance [\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. Pettifor et al. (2000) supported syndromic management but highlighted the need for accessible diagnostics in low-resource settings [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e].\u003c/p\u003e\u003cp\u003e\u003cb\u003eSTUDY AIM\u003c/b\u003e:\u003c/p\u003e\u003cp\u003eDetermine the prevalence of different causative organisms of MUS in South Africa.\u003c/p\u003e\u003cp\u003e\u003cb\u003eSTUDY OBJECTIVES\u003c/b\u003e:\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003ePrimary objective\u003c/span\u003e:\u003c/p\u003e\u003cp\u003e\u003col\u003e\u003cspan\u003e\u003cli\u003e\u003cp\u003eTo determine the prevalence of different causative organisms found on male urethral swabs in patients with urethritis in South Africa.\u003c/p\u003e\u003c/li\u003e\u003c/span\u003e\u003c/ol\u003e\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eSecondary objectives\u003c/span\u003e:\u003c/p\u003e\u003cp\u003e\u003col\u003e\u003cspan\u003e\u003cli\u003e\u003cp\u003eTo determine the rates of antimicrobial resistance in pathogens cultured on urethral swabs of male patients with urethritis in South Africa.\u003c/p\u003e\u003c/li\u003e\u003c/span\u003e\u003cspan\u003e\u003cli\u003e\u003cp\u003eTo compare, within patients, pathogen identification using bacterial cultures to NAAT.\u003c/p\u003e\u003c/li\u003e\u003c/span\u003e\u003cspan\u003e\u003cli\u003e\u003cp\u003e To determine the relevance of current existing guidelines on syndromic management of STI\u0026rsquo;s and whether these are appropriate when compared to our findings.\u003c/p\u003e\u003c/li\u003e\u003c/span\u003e\u003c/ol\u003e\u003c/p\u003e"},{"header":"RESEARCH METHODS AND DESIGN","content":"\u003cp\u003e\u003cb\u003eStudy Design\u003c/b\u003e:\u003c/p\u003e\u003cp\u003eThis was a retrospective cross-sectional study analysing data from 28 July 2012 to 31 December 2023 to describe the microbial aetiology of male urethritis and evaluate antimicrobial resistance patterns.\u003c/p\u003e\u003cp\u003e\u003cb\u003eStudy Setting\u003c/b\u003e:\u003c/p\u003e\u003cp\u003eData were sourced from the central database of Lancet Laboratories, a private diagnostic laboratory network in South Africa.\u003c/p\u003e\u003cp\u003e\u003cb\u003eStudy population and sampling strategy\u003c/b\u003e:\u003c/p\u003e\u003cp\u003eInclusion criteria: Males aged 16 years or older with MUS who submitted urethral swab or first catch urine specimens for NAAT and/or MC\u0026amp;S testing.\u003c/p\u003e\u003cp\u003eSamples were collected and submitted by healthcare workers at various levels of care. All specimens were processed according to WHO-recommended protocols for STI diagnosis using molecular methods [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e] and Clinical and Laboratory Standards Institute protocols [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eLancet laboratories were kind enough to supply anonymised categorised data for the above inclusion criteria in the form of a spreadsheet.\u003c/p\u003e\u003cdiv id=\"Sec2\" class=\"Section2\"\u003e\u003ch2\u003eData Analysis:\u003c/h2\u003e\u003cp\u003eFive datasets were derived after cleaning and verification:\u003c/p\u003e\u003cp\u003e\u003cul\u003e\u003cli\u003e\u003cp\u003eDataset 1: Culture-only results (n\u0026thinsp;=\u0026thinsp;5 021)\u003c/p\u003e\u003c/li\u003e\u003cli\u003e\u003cp\u003eDataset 2: NAAT panel (six organisms) (n\u0026thinsp;=\u0026thinsp;1 055)\u003c/p\u003e\u003c/li\u003e\u003cli\u003e\u003cp\u003eDataset 3: NAAT for \u003cem\u003eN. gonorrhoeae\u003c/em\u003e only (n\u0026thinsp;=\u0026thinsp;27 336)\u003c/p\u003e\u003c/li\u003e\u003cli\u003e\u003cp\u003eDataset 4: NAAT for \u003cem\u003eC. trachomatis\u003c/em\u003e only (n\u0026thinsp;=\u0026thinsp;27 368)\u003c/p\u003e\u003c/li\u003e\u003cli\u003e\u003cp\u003eDataset 5: Paired NAAT and culture data for \u003cem\u003eN. gonorrhoeae\u003c/em\u003e (n\u0026thinsp;=\u0026thinsp;191)\u003c/p\u003e\u003c/li\u003e\u003c/ul\u003e\u003c/p\u003e\u003cp\u003eData were cleaned to remove duplicates and incomplete entries.\u003c/p\u003e\u003cp\u003eAll analyses were conducted in R version 4.4.2. Data were summarised using:\u003c/p\u003e\u003cp\u003e\u003cul\u003e\u003cli\u003e\u003cp\u003eCategorical variables (e.g., organism, resistance status): Frequency and percentage\u003c/p\u003e\u003c/li\u003e\u003cli\u003e\u003cp\u003eContinuous variables (e.g., age): Median, interquartile range (IQR), minimum and maximum\u003c/p\u003e\u003c/li\u003e\u003c/ul\u003e\u003c/p\u003e\u003cp\u003eTo compare NAAT and culture detection of \u003cem\u003eNG\u003c/em\u003e:\u003c/p\u003e\u003cp\u003e\u003cul\u003e\u003cli\u003e\u003cp\u003eSensitivity, specificity, and accuracy of culture were calculated using NAAT as the reference standard\u003c/p\u003e\u003c/li\u003e\u003cli\u003e\u003cp\u003eCohen\u0026rsquo;s Kappa Coefficient was used to assess method agreement [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e].\u003c/p\u003e\u003c/li\u003e\u003c/ul\u003e\u003c/p\u003e\u003cp\u003eComparative resistance data between provinces and organism groups were analysed using Chi-squared tests.\u003c/p\u003e\u003cp\u003e\u003cb\u003eEthical Considerations\u003c/b\u003e:\u003c/p\u003e\u003cp\u003e The study was approved by the Human Research Ethics Committee of the University of the Witwatersrand (medical) (Appendix 1), with data access granted by the Lancet Laboratories Publication Committee (Appendix 2). No patient identifiers were provided, with all data being anonymised at source.\u003c/p\u003e\u003cp\u003eClearance certificate number: M240439\u003c/p\u003e\u003c/div\u003e"},{"header":"RESULTS","content":"\u003cp\u003e\u003cem\u003eDescription of datasets\u003c/em\u003e\u003c/p\u003e\u003cp\u003eAfter cleaning, four sets of data were used in the analysis:\u003c/p\u003e\u003cp\u003e\u003cb\u003eDataset 1 (microbiology only)\u003c/b\u003e (primary objective \u0026amp; secondary objective 1): 5 021 individuals with urethral/penile/meatal swab cultures from 28 July 2012 to 29 Dec 2022. The age range was 16 to 97 years with median age being 37 years. 78.4% (n\u0026thinsp;=\u0026thinsp;3 934) of samples were from Gauteng, and 15.5% from KZN (n\u0026thinsp;=\u0026thinsp;781). Remaining samples were from: Free State: n\u0026thinsp;=\u0026thinsp;52, Mpumalanga: n\u0026thinsp;=\u0026thinsp;151, North West: n\u0026thinsp;=\u0026thinsp;6, Northern Cape: n\u0026thinsp;=\u0026thinsp;32, and Western Cape: n\u0026thinsp;=\u0026thinsp;65.\u003c/p\u003e\u003cp\u003e\u003cb\u003eDataset 2 (NAAT: Six bacterial species)\u003c/b\u003e (primary objective): 1 055 individuals with complete NAAT data for \u003cem\u003eNG, Chlamydia trachomatis, Mycoplasma genitalium, Mycoplasma hominis, Trichomonas vaginalis, Ureaplasma parvum\u003c/em\u003e from urine (n\u0026thinsp;=\u0026thinsp;953) or swab samples (n\u0026thinsp;=\u0026thinsp;102) from 06 May 2015 to 28 Dec 2022. The age range was 16 to 84 years with a median of 35 years. Ninety seven percent (n\u0026thinsp;=\u0026thinsp;1 027) of the samples were from Gauteng. Remaining samples were from: KZN\u0026thinsp;=\u0026thinsp;7, Mpumalanga: n\u0026thinsp;=\u0026thinsp;3, North West: n\u0026thinsp;=\u0026thinsp;1, Northern Cape: n\u0026thinsp;=\u0026thinsp;1, and Western Cape: n\u0026thinsp;=\u0026thinsp;16.\u003c/p\u003e\u003cp\u003e\u003cb\u003eDataset 3 (NAAT\u003c/b\u003e: \u003cb\u003eNG\u003c/b\u003e \u003cb\u003eonly)\u003c/b\u003e (primary objective): 27 366 individuals with NAAT data for \u003cem\u003eNG\u003c/em\u003e DNA from urine samples (n\u0026thinsp;=\u0026thinsp;26 982) or swab samples (n\u0026thinsp;=\u0026thinsp;384) from 17 Dec 2014 to 31 Dec 2022 (8 years). The age range was 16 to 90 years with a median of 35. Eighty five percent (n\u0026thinsp;=\u0026thinsp;23 210) of samples were from Gauteng, 8%, were from KZN (n\u0026thinsp;=\u0026thinsp;2 281), and remaining samples from: Eastern Cape: n\u0026thinsp;=\u0026thinsp;1, Free State: n\u0026thinsp;=\u0026thinsp;288, Mpumalanga: n\u0026thinsp;=\u0026thinsp;585, North West: n\u0026thinsp;=\u0026thinsp;260, Northern Cape: n\u0026thinsp;=\u0026thinsp;103, and Western Cape: n\u0026thinsp;=\u0026thinsp;638.\u003c/p\u003e\u003cp\u003e\u003cb\u003eDataset 4 (NAAT: Assessed for\u003c/b\u003e \u003cb\u003eChlamydia trachomatis\u003c/b\u003e \u003cb\u003eonly)\u003c/b\u003e (primary objective) 27 368 individuals with NAAT data for \u003cem\u003eChlamydia trachomatis\u003c/em\u003e from urine samples (n\u0026thinsp;=\u0026thinsp;26 984) or swab samples (n\u0026thinsp;=\u0026thinsp;384) from 17 Dec 2014 to 31 Dec 2022 (8 years). The age range was 6 to 90 years with a median of 35. Eighty five percent (n\u0026thinsp;=\u0026thinsp;23 211) were from Gauteng, 8.3% were from KZN (n\u0026thinsp;=\u0026thinsp;2 281) with remaining samples from: Eastern Cape: n\u0026thinsp;=\u0026thinsp;1, Free State: n\u0026thinsp;=\u0026thinsp;288, Mpumalanga: n\u0026thinsp;=\u0026thinsp;585, North West: n\u0026thinsp;=\u0026thinsp;260, Northern Cape: n\u0026thinsp;=\u0026thinsp;103, and Western Cape: n\u0026thinsp;=\u0026thinsp;639.\u003c/p\u003e\u003cp\u003e\u003cb\u003eDataset 5 (combined culture and NAAT data)\u003c/b\u003e (secondary objective 2): 191 individuals with both NAAT and culture data for \u003cem\u003eNG\u003c/em\u003e from 27 September 2013 to 12 December 2022 (~\u0026thinsp;9 years). The age range was 17 to 77 years with a median of 34. Eighty six percent (n\u0026thinsp;=\u0026thinsp;165) came from Gauteng with remaining samples from: KZN: n\u0026thinsp;=\u0026thinsp;20, Mpumalanga: n\u0026thinsp;=\u0026thinsp;2, and Western Cape: n\u0026thinsp;=\u0026thinsp;4. ~35% of NAAT assays detected \u003cem\u003eNG\u003c/em\u003e, and ~\u0026thinsp;19% of cultures detected \u003cem\u003eNG\u003c/em\u003e.\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003ePrimary objective\u003c/span\u003e:\u003c/p\u003e\u003cp\u003eTo determine the prevalence of different causative organisms in MUS in South Africa using bacterial culture and NAAT.\u003c/p\u003e\u003cp\u003eDataset 1:\u003c/p\u003e\u003cp\u003e5,021 cultures (2012\u0026ndash;2022); 58% showed no growth. NG was most frequent (12%), followed by Candida and E. coli.\u003c/p\u003e\u003cp\u003eIn 5,021 cultures, 58% were negative; 42% yielded organisms. NG was most common (12%).\u003c/p\u003e\u003cp\u003eNeisseria gonorrhoeae (NG) was the most frequently isolated organism (~\u0026thinsp;12%), occurring 3.4 times more often than the next most common, Candida. Escherichia coli was the most common bacterial species after NG (3.3%). Ureaplasma urealyticum, a recognised STI pathogen, was the sixth most common isolate (2%).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eDataset 2:\u003c/p\u003e\u003cp\u003e1,055 NAATs for six species (2015\u0026ndash;2022); 80% were negative. CT, UP, and MG were most common positives.\u003c/p\u003e\u003cp\u003eOf 1,055 NAAT samples, NG was 2.1%. Most positives were non-gonococcal.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eDataset 3:\u003c/p\u003e\u003cp\u003e27,336 NAATs for NG (2014\u0026ndash;2022); 3.8% positive.\u003c/p\u003e\u003cp\u003eAmong 27,336 NAATs for NG, 3.8% were positive.\u003c/p\u003e\u003cp\u003eDataset 4:\u003c/p\u003e\u003cp\u003e27,368 NAATs for CT (2014\u0026ndash;2022); 7.1% positive.\u003c/p\u003e\u003cp\u003eOf 27,368 tested for CT, 7.1% were positive.\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eSecondary objective 1\u003c/span\u003e:\u003c/p\u003e\u003cp\u003eDetermine the frequency of antimicrobial resistance in STI pathogens cultured from urethral swabs of male patients with urethritis in South Africa.\u003c/p\u003e\u003cp\u003eDetermine the frequency of antimicrobial resistance in STI pathogens cultured from urethral swabs of male patients with urethritis in South Africa.\u003c/p\u003e\u003cp\u003eNeisseria gonorrhoeae showed high sensitivity to azithromycin (89.3%) and cefixime (93.8%), but markedly lower rates for ciprofloxacin (34.9%), penicillin (23.9%), and tetracycline (13.1%) (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eUreaplasma urealyticum demonstrated moderate sensitivity to erythromycin (75.5%) and tetracycline (61.2%) (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eProvincial variation in NG sensitivity (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003e):\u003c/p\u003e\u003cp\u003eAntibiotic sensitivity data were analysed for Gauteng, KZN, and Western Cape (\u0026ge;\u0026thinsp;10 NG cultures per antibiotic). Azithromycin data were only available from Gauteng; KZN lacked tetracycline data.\u003c/p\u003e\u003cp\u003eCefixime sensitivity was high in Gauteng (94%) and KZN (95.2%), but lower in WC (78.6%). Ciprofloxacin susceptibility was poor overall but highest in KZN (51.2%) compared to Gauteng (27.9%) and WC (38.5%). Penicillin susceptibility followed a similar trend: KZN (29.5%)\u0026thinsp;\u0026gt;\u0026thinsp;Gauteng (17.6%)\u0026thinsp;\u0026gt;\u0026thinsp;WC (14.2%). Tetracycline resistance was high in both Gauteng (88.1%) and WC (54.5%).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eLimitations and combination therapy analysis (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003e):\u003c/p\u003e\u003cp\u003eOnly Gauteng had sufficient Ureaplasma urealyticum data for analysis, limiting interprovincial comparison.\u003c/p\u003e\u003cp\u003eCurrent first-line syndromic management includes ceftriaxone, azithromycin, and doxycycline. Doxycycline was not tested against NG and was used in only two U. urealyticum cultures\u0026mdash;each as the sole antibiotic tested. No NG cultures included ceftriaxone, but azithromycin and cefixime were assessed.\u003c/p\u003e\u003cp\u003eAmong NG isolates with both results, 84% were sensitive to both drugs, 9.7% were cefixime-sensitive but azithromycin-resistant, 5.2% showed the reverse, and dual resistance was rare (1.1%).\u003c/p\u003e\u003cp\u003eOnly Gauteng and WC contributed to this analysis, with just four cases from WC, precluding meaningful provincial comparison.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003eSecondary objective 2: Culture vs NAAT for NG detection (\u003c/span\u003eFig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003e\u003cspan type=\"Underline\" class=\"Underline\" name=\"Emphasis\"\u003e)\u003c/span\u003e:\u003c/p\u003e\u003cp\u003eA total of 191 patients had matched NAAT and culture data for Neisseria gonorrhoeae. Using NAAT as the reference standard, culture showed high specificity but limited sensitivity.\u003c/p\u003e\u003cp\u003eCulture accuracy was 87.4% (95% CI: 81.9\u0026ndash;91.8), sensitivity 56%, and specificity 98.6%. Cohen\u0026rsquo;s Kappa was 0.63, indicating moderate-to-good agreement between methods.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eCulture was negative in 84.3% (n\u0026thinsp;=\u0026thinsp;161) and positive in 15.7% (n\u0026thinsp;=\u0026thinsp;30). NAAT detected NG in 50 cases (26.2%), but 22 of these were missed by culture. Only two samples were culture-positive but NAAT-negative, highlighting a high false-negative rate for culture.\u003c/p\u003e"},{"header":"DISCUSSION","content":"\u003cp\u003eThe study provides a detailed overview of aetiology and resistance patterns of sexually transmitted MUS in South Africa\u0026rsquo;s private healthcare sector over a ten-year period. The study incorporates both culture-based and molecular diagnostic techniques, resulting in one of the largest and most comprehensive datasets of its kind in South Africa. The findings have important implications for current syndromic management algorithms and diagnostic modalities.\u003c/p\u003e\u003cp\u003e\u003cb\u003ePathogen Distribution\u003c/b\u003e:\u003c/p\u003e\u003cp\u003eNG was the most commonly identified pathogen (~\u0026thinsp;12%). Urine is less invasive and easier to handle than swabs, influencing clinician preference. These findings support molecular diagnostics for NGU screening.\u003c/p\u003e\u003cp\u003e\u003cb\u003eAntimicrobial Resistance Patterns\u003c/b\u003e:\u003c/p\u003e\u003cp\u003e\u003cb\u003eNeisseria gonorrhoeae\u003c/b\u003e\u003c/p\u003e\u003cp\u003eThe AMR profile of NG demonstrated marked resistance variability. Lowest resistance was observed for cefixime (5.8%) and azithromycin (10.7%), while ciprofloxacin (65.1%), penicillin (76.1%), and tetracycline (86.9%) showed unacceptably high resistance. Quinolone resistance rose sharply from 0\u0026ndash;22% in KZN in 2023 to 65.1% [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e], echoing global trends and reinforcing the exclusion of ciprofloxacin and tetracycline from empiric regimens [\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e, \u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e]. These trends align with the WHO\u0026rsquo;s classification of NG AMR as a high-priority threat [\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eRegional differences were notable: Cefixime susceptibility in WC was 78.6%, compared to 94% in Gauteng and 95.2% in KZN, suggesting regional clonal variation or prescribing differences. Prior AMR data were limited to Gauteng, KZN, Eastern Cape, and WC [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e], yet resistance appears to be worsening.\u003c/p\u003e\u003cp\u003eTetracycline-resistant NG (TRNG) is now globally prevalent [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e]. In South Africa, TRNG increased from 73% in 2011 to 86.9% in the present data [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e]. Prior studies noted a preference for oral cephalosporins (cefixime) in high-income settings versus intramuscular ceftriaxone in lower-income regions [\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e]. This distinction is evident locally: Cefixime use dominates in the private sector, likely due to ease of oral administration and less price sensitivity, whereas ceftriaxone remains standard in the public sector.\u003c/p\u003e\u003cp\u003eDespite cefixime not being recommended in MUS guidelines or listed on the government\u0026rsquo;s essential medicines list [\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e], its use persists in the private sector, even with greater resistance than ceftriaxone [\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e]. Private providers are thus advised to align with syndromic protocols despite convenience or patient preference for oral therapy.\u003c/p\u003e\u003cp\u003e\u003cb\u003eComparison to Previous Studies on Neisseria gonorrhoeae in South Africa\u003c/b\u003e\u003c/p\u003e\u003cp\u003eThe AMR patterns largely reflect, but also diverge from, those reported by Yakobi et al. (2024). As similarly observed, high resistance to ciprofloxacin, penicillin, and tetracycline was found across Gauteng, KZN, and WC. However, ciprofloxacin resistance was significantly higher in the current study (65.1%) compared to Yakobi et al.\u0026rsquo;s findings (23% in KZN, 14% in Gauteng, 100% in WC), possibly due to evolving resistance or private sector prescribing patterns [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e].\u003c/p\u003e\u003cp\u003ePenicillin susceptibility was low (23.9%), aligning with resistance reported in Gauteng (23%) and KZN (25%) by Yakobi et al. [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. In WC, susceptibility was even lower (14.2%), although Yakobi et al. did not provide comparative data for this province.\u003c/p\u003e\u003cp\u003eTetracycline resistance was extensive (86.9%), exceeding Yakobi et al.\u0026rsquo;s national rate (30%) (5). Gauteng showed 88.1% resistance, WC 54.5%, compared to Yakobi\u0026rsquo;s 61% in Gauteng and 36% in KZN [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eIn contrast, azithromycin (89.3%) and cefixime (93.8%) retained high susceptibility, consistent with Yakobi et al.\u0026rsquo;s low resistance rates (azithromycin: 17% in KZN, 1% in Gauteng; cefixime: \u0026le;0.6%) [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. However, cefixime susceptibility was lower in WC (78.6%) than in Gauteng and KZN (94\u0026ndash;95%), warranting surveillance.\u003c/p\u003e\u003cp\u003e Dual susceptibility to azithromycin and cefixime was found in 84% of isolates, supporting current treatment guidelines. Notably, this is the first report of XDR N. gonorrhoeae in South Africa [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e], with a prevalence of 1.1%. XDR NG is defined as resistance to both recommended therapies (cephalosporin and azithromycin) plus resistance to two or more other antimicrobials (penicillin, tetracycline, erythromycin) [\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e]. Among the three XDR cases, one isolate was sensitive to penicillin, one to tetracycline, and one resistant to all five agents tested.\u003c/p\u003e\u003cp\u003eThese findings highlight provincial AMR differences, underscoring the need for regional surveillance.\u003c/p\u003e\u003cp\u003e\u003cb\u003eUreaplasma urealyticum\u003c/b\u003e:\u003c/p\u003e\u003cp\u003eResistance in Ureaplasma urealyticum (UU) is notable due to its lack of a cell wall, rendering it naturally resistant to all cell-wall-targeting antibiotics, particularly β-lactams [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e], which limits treatment options and exacerbates resistance concerns.\u003c/p\u003e\u003cp\u003eResistance to first-line macrolides and tetracyclines was prominent, especially tetracycline (38.8%), while susceptibility to erythromycin remained relatively preserved at 75.5% [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e]. This resistance markedly exceeds global averages reported by Ramaloko et al. (2025), where macrolide and tetracycline resistance were 19% and 8%, respectively [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e]. The observed tetracycline resistance is roughly fivefold higher than global estimates (38.8% vs 8%), raising concerns about empiric doxycycline use in syndromic STI management [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e, \u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eSimilarly, the 24.5% erythromycin resistance rate surpasses the global mean of 19%, indicating worsening macrolide resistance in local Ureaplasma strains, potentially driven by widespread azithromycin use in syndromic protocols [\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e, \u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e]. Historically, UU has been considered more macrolide-susceptible than Mycoplasma genitalium, but this study suggests this may no longer hold true in South African private healthcare settings.\u003c/p\u003e\u003cp\u003eWhile fluoroquinolone resistance and multidrug resistance (MDR) were not assessed here, these remain critical areas for future surveillance. Given high resistance rates to two primary antimicrobial classes, MDR UU may be circulating undetected in South Africa.\u003c/p\u003e\u003cp\u003eAntimicrobial resistance studies remain limited nationally [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]. Provinces such as Mpumalanga and North West have never undergone NG AMR surveillance, and many regions lack NG susceptibility data since the introduction of syndromic management in the 1990s [\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e]. Enhanced routine STI aetiological monitoring is urgently needed to update syndromic treatment regimens [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e], despite challenges posed by budget and logistical constraints. This is vital for South Africa\u0026rsquo;s progress toward the WHO target of 90% STI reduction by 2030 [\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e, \u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e]. Europe\u0026rsquo;s modest 3% annual decline in GU incidence since this target was set illustrates the difficulty of this task [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e].\u003c/p\u003e\u003cp\u003e\u003cb\u003eComparison of Culture\u003c/b\u003e \u003cb\u003evs\u003c/b\u003e \u003cb\u003eNAAT\u003c/b\u003e:\u003c/p\u003e\u003cp\u003eIn 191 samples tested for Neisseria gonorrhoeae (NG) by both NAAT and culture, culture demonstrated a sensitivity of 56% and specificity of 98.6%, with Cohen\u0026rsquo;s κ\u0026thinsp;=\u0026thinsp;0.63, indicating good agreement. In contrast, Serra-Pladevall et al. (2015) reported higher culture sensitivity (86.2%) and specificity (99.8%) [\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e], their study treated urine specimens as equivalent to urethral swabs, differing from the present methodology [\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e]. NAAT in their study showed sensitivity and specificity of 98.7% and 100% respectively [\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eThus, while specificities are comparable, the markedly lower culture sensitivity here suggests up to 44% of NG infections may be missed. These findings confirm culture\u0026rsquo;s known limitations as a low-sensitivity, high-specificity diagnostic tool for NG [\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e]. The moderate agreement and poor sensitivity highlight that culture alone is insufficient and may lead to under-treatment.\u003c/p\u003e\u003cp\u003e\u003cb\u003eStrengths and Limitations of Study\u003c/b\u003e:\u003c/p\u003e\u003cp\u003eThis is the first large-scale private sector analysis of MUS pathogens and resistance in South Africa.\u003c/p\u003e\u003cp\u003eLimitations include reliance solely on laboratory results without access to specimen collection methods, patient history, or clinical symptoms. Standardisation across this large, geographically diverse cohort was not feasible. Additionally, the study lacked ceftriaxone resistance data, may be affected by selection bias, and did not include longitudinal follow-up or treatment outcomes.\u003c/p\u003e"},{"header":"Conclusions","content":"\u003cp\u003eThe study supports ongoing dual therapy for MUS in South Africa but raises concerns about the long-term use of azithromycin due to emerging resistance. It also advises against routine doxycycline use in syndromic management owing to high resistance rates.\u003c/p\u003e\u003cp\u003eThe study highlights the limitations of culture in diagnosing NGU and underscores the increasing value of NAAT-based multiplex panels, especially in atypical or persistent cases. However, culture remains essential for antimicrobial resistance profiling.\u003c/p\u003e\u003cp\u003eThe unprecedented finding of XDR NG in South Africa is of great concern to public health.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003eThe authors have no financial or personal relationships that may have inappropriately influenced the content of this thesis. No conflicts of interest are present.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors have no conflicts of interest to declare that are relevant to the content of this article.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNo funding was received for conducting this study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDisclaimer:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe views expressed in this article are those of the author and not of any associated institution.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions:\u003c/strong\u003e\u003c/p\u003e\n\u003cul\u003e\n \u003cli\u003eDr. Matthew Snyman is the corresponding author for this study, and conducted all communication with outside parties, conducted the literature review, collected and analysed data, drew conclusions and wrote the final paper as an original work.\u003c/li\u003e\n \u003cli\u003eDr. Marietha Nel and Dr. Winston Padayachee jointly supervised this study in equal contribution, providing valuable insight, advice and assistance in proofreading and correcting the final write-up.\u003c/li\u003e\n\u003c/ul\u003e\n\u003cp\u003e\u003cstrong\u003eDEDICATION:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo my family, for their unwavering support, to my mentors, who guided me and to my patients who have inspired me.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eETHICS APPROVAL:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eUniversity of the Witwatersrand Human Research Ethics Committee (Medical) approved this study and designated M240439 as a reference number\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eACKNOWLEDGEMENTS:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eDr Marietha Nel and Dr Winston Padayachee, for your supervision and guidance overseeing my execution of this research and write-up.\u003c/p\u003e\n\u003cp\u003eProf Peter Kamerman for assistance with the statistics and data analysis of results and for your invaluable insights and guidance.\u003c/p\u003e\n\u003cp\u003eProfessor Ahmed Adam for your guidance and expert advice\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eLancet Laboratories for kindly providing access to their extensive database of results\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\u003cli\u003e\u003cspan\u003eEAU Guidelines Office EAU Guidelines. 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Diagn Microbiol Infect Dis 83(4):341\u0026ndash;343\u003c/span\u003e\u003c/li\u003e\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":true,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"world-journal-of-urology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"wjur","sideBox":"Learn more about [World Journal of Urology](https://link.springer.com/journal/345)","snPcode":"345","submissionUrl":"https://submission.nature.com/new-submission/345/3","title":"World Journal of Urology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false},"keywords":"Urethritis, Sexually transmitted infection, Incidence, Resistance, Chlamydia trachomatis, Neisseria Gonorrhoea ","lastPublishedDoi":"10.21203/rs.3.rs-7085306/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7085306/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003e\u003cstrong\u003ePurpose:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo assess the microbial aetiology and AMR patterns in MUS by analysing private laboratory urethral swab and urine specimens. The study also compared diagnostic performance between NAAT and culture and evaluated the relevance of syndromic management. This is the first large-scale South African private sector analysis of MUS pathogen profiles and AMR using NAAT and culture, providing critical evidence for diagnostic and management policy updates.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eMethods:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eA retrospective cross-sectional study was performed using laboratory data from 2012 to 2023, comprising over 61,000 samples. Organism prevalence and AMR rates were assessed. Diagnostic sensitivity and specificity of NAAT versus culture for Neisseria gonorrhoeae (\u003cem\u003eNG\u003c/em\u003e) were calculated in matched samples.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eResults:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cem\u003eNG\u003c/em\u003e and \u003cem\u003eChlamydia trachomatis\u003c/em\u003e were the most prevalent pathogens. NAAT demonstrated superior sensitivity, while culture yielded essential AMR data. \u003cem\u003eNG\u003c/em\u003e showed elevated resistance patterns, but susceptibility to first-line antimicrobials remained high. Culture sensitivity and specificity relative to NAAT were 56% and 98.6%, respectively. Ureaplasma urealyticum showed elevated resistance versus global averages. The first cases of extensively drug-resistant (XDR) NG in South Africa were identified.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConclusion:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNAAT improves MUS detection, while culture remains vital for AMR surveillance. Syndromic management remains effective, but rising resistance and XDR NG emergence warrant urgent attention.\u003c/p\u003e","manuscriptTitle":"Distribution and resistance patterns of male urethritis pathogens from a private South African laboratory","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-08-04 08:41:28","doi":"10.21203/rs.3.rs-7085306/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2025-10-29T14:06:37+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2025-10-24T09:27:14+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"120584335317385941230463680128408536095","date":"2025-10-06T09:17:26+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2025-07-30T09:25:11+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2025-07-22T03:57:47+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2025-07-21T16:34:38+00:00","index":"","fulltext":""},{"type":"submitted","content":"World Journal of Urology","date":"2025-07-09T15:14:45+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"world-journal-of-urology","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"wjur","sideBox":"Learn more about [World Journal of Urology](https://link.springer.com/journal/345)","snPcode":"345","submissionUrl":"https://submission.nature.com/new-submission/345/3","title":"World Journal of Urology","twitterHandle":"","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"stoa","reportingPortfolio":"Springer Hybrid","inReviewEnabled":true,"inReviewRevisionsEnabled":false}}],"origin":"","ownerIdentity":"c4548ca6-111f-43f8-9d84-1e70e7702b5c","owner":[],"postedDate":"August 4th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"published-in-journal","subjectAreas":[],"tags":[],"updatedAt":"2025-12-15T16:00:32+00:00","versionOfRecord":{"articleIdentity":"rs-7085306","link":"https://doi.org/10.1007/s00345-025-06117-y","journal":{"identity":"world-journal-of-urology","isVorOnly":false,"title":"World Journal of Urology"},"publishedOn":"2025-12-11 15:57:00","publishedOnDateReadable":"December 11th, 2025"},"versionCreatedAt":"2025-08-04 08:41:28","video":"","vorDoi":"10.1007/s00345-025-06117-y","vorDoiUrl":"https://doi.org/10.1007/s00345-025-06117-y","workflowStages":[]},"version":"v1","identity":"rs-7085306","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7085306","identity":"rs-7085306","version":["v1"]},"buildId":"8U1c8b4HqxoKbykW_rLl7","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}