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Introduction: Home-based self-collected dried blood spot (DBS) sampling could simplify sexual health and preexposure prophylaxis care and reduce sexually transmitted infections (STIs) clinic visits for men who have sex with men (MSM). We compared the performance of DBS to venipuncture collected blood samples to test four STIs and creatinine concentration., Methods: We invited MSM clients of the Amsterdam STI clinic to participate. Routinely collected peripheral blood was tested for syphilis treponemal antibody, HIV (HIV Ag/Ab), HCV (antibodies), HBV (HBsAg) and creatinine concentration. Participants received a home kit for DBS sampling, a return envelope and a questionnaire to evaluate the acceptability, feasibility and usability of DBS, measured on 5-point Likert scales, 1 representing complete disagreement and 5 complete agreement. We assessed sensitivity and specificity of DBS versus peripheral blood-based testing., Results: In 2020 to 2021, we included 410 participants; 211 (51.5%) returned a completed DBS card, 117 (28.5%) returned a partially filled card and 82 (20.0%) did not return a card. The sensitivity for syphilis was 90.8% and the specificity 84.3%. For both HIV Ag/Ab and HBsAg, the sensitivity and specificity were 100.0%. The sensitivity for HCV antibody was 80.0%, and the specificity was 99.2%. The DBS creatinine concentration was a mean of 5.3 μmol/L higher than in venipuncture obtained plasma. Participants' median willingness to take a future DBS was 4 (interquartile range, 3-5)., Discussion: Dried blood spot may be an acceptable method among MSM for STI testing and creatinine follow-up during preexposure prophylaxis use. However, collecting enough blood on DBS cards was a challenge, and sensitivities for syphilis and HCV serology were too low., Competing Interests: Conflict of Interest and Sources of Funding: None declared., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Sexually Transmitted Diseases Association.)

Background: Syphilis is a complex, multistage, sexually transmitted infection (STI) caused by the bacterium Treponema pallidum subspecies pallidum (TP). New diagnostic tools are needed to minimize transmission. In this study, we aimed to assess the additional value of an investigational transcription-mediated amplification test for TP (TP-TMA) for routine diagnostics., Methods: Between September 2021 and August 2022, visits by all participants of the national preexposure prophylaxis (PrEP) program at the sexual health center (SHC) in Amsterdam were included. Anal, pharyngeal, vaginal, and urine samples collected for Chlamydia trachomatis and Neisseria gonorrhoeae screening were additionally tested with the TP-TMA assay based on detection of 23S rRNA of TP., Results: In total, 9974 SHC visits by 3283 participants were included. There were 191 infectious syphilis cases diagnosed: 26 (14%) primary syphilis, 54 (29%) secondary syphilis, and 111 (58%) early latent syphilis. In 79 of the 191 (41%) syphilis cases, at least 1 sample was TP-TMA-positive. For 16 participants, the positive TP-TMA result was not concordant with routine diagnostics. Of those, 2 participants were treated for syphilis within a week before the visit. Eight participants were treated for a syphilis notification at the visit or for another STI. Five participants were diagnosed with syphilis at the following visit, and 1 participant was lost to follow-up., Conclusions: By adding the TP-TMA assay to routine diagnostics, we identified 14 of 191 (7%) additional syphilis infections among participants of the national PrEP program. The TP-TMA assay is a useful diagnostic tool to increase syphilis case finding and thus limit the transmission of syphilis., Competing Interests: Potential conflicts of interest. D. G. reports financial or nonfinancial interests as an employee of Hologic and a patent planned, issued, or pending related to detection of TP. A. N. reports employment with and stock with Hologic. All other authors report no potential conflicts. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed., (© The Author(s) 2023. Published by Oxford University Press on behalf of Infectious Diseases Society of America.)

Background: Diagnosis of infected individuals with Mycoplasma genitalium (MG) is often performed by real-time PCR or transcription-mediated amplification (TMA). A limitation of the MG-TMA assay is the relatively short time span of 24 h in which the collected urine is required to be transferred into a Urine Specimen Transport Tube, according to the manufacturer's guidelines. If not transferred within 24 h, the manufacturer's claimed sensitivity cannot be guaranteed anymore, and samples may instead be tested with an in-house validated real-time PCR, despite its recognized lower sensitivity. This study aimed to validate an exception to the sample transport and storage conditions of the MG-TMA assay as set by the manufacturer, being the prolongation of the acceptable testing time limit of 24 h., Methods: From June to December 2022, first-void urines were collected from clients attending the clinic for sexual health in Amsterdam, the Netherlands. Urine samples that tested positive for MG by TMA assay at the day of collection were concomitantly stored at room (18-24 °C) and refrigerator temperature (4-8 °C) for 15 days. The stored urine samples were tested with both an in-house validated real-time PCR and MG-TMA assay after transfer of the original urine samples to the respective test tubes at 3, 7, 12 and 15 days post collection., Results: In total, 47 MG-positive urine samples were collected, stored and tested for MG by real-time PCR and TMA assays. After storage at room temperature, the MG-detection rate by TMA was significantly higher compared to real-time PCR, at days 0 (p ≤ 0.001), 7 (p ≤ 0.001) and 12 (p < 0.05). After storage at refrigerator temperature, the MG-detection rate determined by TMA assay was significantly enhanced in comparison with real-time PCR at days 3 (p < 0.01), 7 (p ≤ 0.001) and 15 (p < 0.01)., Conclusions: This validation study showed that the MG-TMA assay has a superior detection rate in urine compared to real-time PCR, up to 15 days post sample collection and irrespective of storage temperature. Accepting urines older than 24 h to be tested by TMA will improve clinical diagnosis of MG infections., (© 2023. BioMed Central Ltd., part of Springer Nature.)

Background: People experiencing homelessness (PEH) may be at increased risk of SARS-CoV-2 infection and severe COVID-19. The Dutch government established emergency shelters and introduced preventive measures for homelessness services. There were no major SARS-CoV-2 outbreaks noticed among PEH during the first two waves of infections. This study aimed to assess the prevalence of current and past infections among PEH and staff by conducting an on-site COVID-19 screening project at homelessness services in Amsterdam, the Netherlands., Methods: We assessed the proportion of visitors and staff members of four homelessness services at two locations in Amsterdam with positive SARS-CoV-2 qPCR and antibody results (IgG/IgM Rapid Test/Biozek) in May 2021. We also assessed sociodemographic, clinical and lifestyle characteristics, compliance with basic prevention measures and intention to vaccinate against COVID-19 among PEH and staff., Results: A total of 138 visitors and 53 staff members filled out a questionnaire and were tested. Among PEH, the SARS-CoV-2 positivity rate was 0% (0/133;95%CI = 0-1.9) and the antibody positivity rate was 1.6% (2/131;95%CI = 0.8-7.5) among those without prior COVID-19 vaccination. Among staff, these percentages were 3% (1/32;95%CI = 0.1-16.2) and 11% (5/53;95%CI = 3.6-23.6), respectively. Most participants were often compliant with the basic preventive measures 'not shaking hands', 'wearing a face mask' and 'washing hands', but not with 'physical distancing'. High vaccination intent was more common among staff members (55%) than among visitors (42%), while high trust in the governmental COVID-19 policies was more common among visitors (41%) than among staff (30%)., Conclusions: We observed a low prevalence of past and current SARS-CoV-2 infections among PEH, which may be explained by instated shelter policies, limited daily activities of PEH and compliance with prevention measures. Vaccine hesitancy and mistrust among visitors and staff could hinder vaccination uptake, suggesting that interventions towards homelessness services are needed., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Generaal et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Objectives: A previous study showed higher prevalence of Escherichia coli (E. coli) expressing extended-spectrum β-lactamases (ESBL-Ec) among men who have sex with men (MSM) in Amsterdam, the Netherlands, compared with the general Dutch population. This study genetically characterised the ESBL-Ec isolates and investigated whether the increased prevalence could be explained by transmission between participants., Methods: Whole-genome sequences were obtained from 93 unique ESBL-Ec isolates that were cultured from rectal swabs of 79 participants. Isolates were typed according to the Achtman MLST scheme and ESBL and virulence genes were identified. Pairwise SNP distances were determined between isolates. Isolate pairs with ≤ 25 SNPs were considered part of a putative transmission event, and events between multiple participants formed putative transmission clusters. To investigate whether putatively transmitted isolates belonged to globally expanded lineages, the level of hierarchical clustering with international isolates was assessed using core genome MLST (cgMLST) implemented on the Enterobase platform., Results: The most frequently detected E. coli types were ST131:bla CTX-M-15 (16 of 117, 13.5%), ST131:bla CTX-M-27 , ST3075:bla CTX-M-15 and ST14:bla SHV-12 (all six of 117, 6.5%). Fourteen putative transmission events were identified, forming four putative transmission clusters. The largest putative transmission cluster contained ST131 isolates, which clustered with multiple international isolates in SNP and cgMLST analysis. One other transmission cluster (ST14:bla SHV-12 ) and two transmission events (ST14:bla SHV-12 and ST394:bla CTX-M-15 ) contained very rarely reported strains., Conclusions: The identification of unique ESBL-Ec strains involved in putative transmission and carried by multiple participants demonstrated a high probability of ESBL-Ec transmission between MSM in Amsterdam; therefore, ESBL-Ec infection should be considered in cases of sexually active MSM having associated symptoms., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: During the first wave of COVID-19 in Amsterdam, the Netherlands, a disproportional number of COVID-19 hospitalizations occurred in individuals with an ethnic minority background and in individuals living in city districts with a lower socioeconomic status (SES). In this study, we assessed whether these disparities continued throughout the second wave, when SARS-CoV-2 testing was available to anyone with symptoms but prior to the availability of COVID-19 vaccination., Methods: Surveillance data on all notified SARS-CoV-2 cases in Amsterdam between 15 June 2020 and 20 January 2021 were matched to municipal registration data to obtain the migration background of cases. Crude and directly age- and sex-standardized rates (DSR) of confirmed cases, hospitalizations, and deaths per 100,000 population were calculated overall, and by city districts, and migration backgrounds. Rate differences (RD) and rate ratios (RR) were calculated to compare DSR between city districts and migration backgrounds. We used multivariable Poisson regression to assess the association of city districts, migration backgrounds, age, and sex with rates of hospitalization., Results: A total of 53,584 SARS-CoV-2 cases (median age 35 years [IQR = 25-74]) were notified, of whom 1,113 (2.1%) were hospitalized and 297 (0.6%) deceased. DSR of notified infections, hospitalization, and deaths per 100,000 population were higher in lower SES peripheral city districts (South-East/North/New-West) than higher SES central districts (Central/West/South/East), with almost a 2-fold higher hospitalization DSR in peripheral compared to central districts (RR = 1.86, 95%CI = 1.74-1.97). Individuals with a non-European migration background also had a higher COVID-19 burden, particularly with respect to hospitalization rates, with a 4.5-fold higher DSR for individuals with a non-European background compared to ethnic-Dutch (RR 4.51, 95%CI = 4.37-4.65). City districts, migration backgrounds, male gender, and older age were independently associated with COVID-19 hospitalization rates., Discussion: Individuals with a non-European background and individuals living in city districts with lower SES continued to independently have the highest COVID-19 burden in the second wave of COVID-19 in Amsterdam, the Netherlands., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Bachour, Wynberg, Coyer, Buster, Schreijer, van Duijnhoven, van Dam, Prins and Leenstra.)

Background: Antibiotic-impregnated cement-coated plates (ACPs) have been used successfully for temporary internal fixation between stages in the two-stage treatment of infected non-unions. We describe our approach of using an ACP in the staged treatment of a methicillin-resistant Staphylococcus aureus (MRSA)-infected distal femoral non-union below a total hip prosthesis. In addition, we present the results of an in vitro experiment to provide an in-depth insight into the capacity of ACPs in (i) treating residual biofilm and (ii) preventing bacterial recolonisation., Materials and Methods: In the first stage, we used a titanium LISS plate coated with hand-mixed PALACOS with vancomycin (PAL-V) for temporary internal fixation combined with commercially prepared COPAL with gentamicin and vancomycin (COP-GV) to fill the segmental defect. In the second stage, the non-union was treated with double-plate fixation and bone grafting.A Kirby-Bauer agar disc diffusion assay was performed to determine the antimicrobial activity of both ACPs and a drug-release assay to measure antibiotic release over time. A biofilm killing assay was also carried out to determine if the antibiotic released was able to reduce or eradicate biofilm of the patient's MRSA strain., Results: At one-year follow-up, there was complete bone-bridging across the previous non-union. The patient was pain-free and ambulatory without need for further surgery. Both ACPs with COP-GV and PAL-V exerted an antimicrobial effect against the MRSA strain with peak concentrations of antibiotic released within the first 24 hours. Concentrations released from COP-GV in the first 24 hours in vitro caused a 7.7-fold log reduction of colony-forming units (CFU) in the biofilm. At day 50, both COP-GV and PAL-V still released concentrations of antibiotic above the respective minimal inhibitory concentrations (MIC), likely contributing to the positive clinical outcome., Conclusion: The use of an ACP provides stability and infection control in the clinical scenario of an infected non-union. This is confirmed in vitro where the release of antibiotics from ACPs is characterised by an early burst followed by a prolonged sustained release above the MIC until 50 days. The burst release from COP-GV reduces CFU in the biofilm and prevents early recolonisation through synergistic activity of the released vancomycin and gentamicin., Clinical Significance: An antibiotic-impregnated cement-coated plate is a useful addition to the surgeon's armamentarium to provide temporary internal fixation without the disadvantages of external fixation and contribute to infection control in an infected non-union., How to Cite This Article: Wagner RK, Guarch-Pérez C, van Dam AP, et al . Antimicrobial Mechanisms and Preparation of Antibiotic-impregnated Cement-coated Locking Plates in the Treatment of Infected Non-unions. Strategies Trauma Limb Reconstr 2023;18(2):73-81., Competing Interests: Source of support: This work was funded by the research project PRINT-AID, within the EU Framework Programme for Research and Innovation Horizon 2020—Marie Sklodowska-Curie Innovative Training Networks under grant agreement no. 722467. Other funding for the conduction of the experiment was departmental. The PALACOS LV cement and a mixing system were provided by Heraeus Nederland BV. The funders had no role in study design, data collection and analysis, decision to publish or preparation of the paper. Conflict of interest: None, (Copyright © 2023; The Author(s).)

Distancing measures during the COVID-19 lockdown led to a temporary decrease of casual sex partners among clients of the Centre for Sexual Health (CSH) in Amsterdam, the Netherlands. We investigated the effect of this change on the genotypic and phenotypic distribution of Neisseria gonorrhoeae ( Ng ) isolates from CSH patients. From each Ng- positive patient we sequenced one isolate, resulting in 322 isolates which constituted two groups: 181 isolates cultured from 15 January to 29 February 2020 (before the first lockdown) and 141 cultured from 15 May to 30 June 2020 (during the first lockdown). Patient characteristics showed significantly more symptomatic patients and significantly fewer reported sex partners during the lockdown. Phenotypic data showed an increase in low-level azithromycin resistance and ceftriaxone susceptibility during the lockdown, and this remained after the study period. The diversity in sequence types (STs) decreased slightly during the lockdown. A shift occurred from ST 8156 being predominant before lockdown to ST 9362 during lockdown and a remarkably low median SNP distance of 17 SNPs was found between ST 9362 isolates obtained during lockdown. These findings reflect restricted travel and the change in sexual behaviour of CSH clients during the lockdown, with a potentially increased local transmission of the ST 9362 strain during this period, which led to genotypic and phenotypic changes in the Ng population. This shows that public health measures have far-reaching consequences and should be considered in the surveillance of other infectious diseases.

Objective: This study aimed to determine the prevalence of fluoroquinolone resistance-associated mutations (QRAMs) in Mycoplasma genitalium (MG) among clients of two sexual health centres (SHCs) in the Netherlands., Design: A cross-sectional study., Setting and Participants: Between 2018 and 2019, 669 clients with MG were included from two previous studies: 375 male clients with urethritis from the SHC in Amsterdam; and 294 clients (male and female) from the SHC in Amsterdam and The Hague. Urogenital and anal samples (705 in total) that tested positive for MG by nucleic acid amplification tests were selected., Outcome Measures: The presence of QRAM was detected by an MG-QRAM PCR targeting four mutations in the parC gene and investigated by sequence analysis of relevant regions of the gyrA and parC genes. Possible risk factors for the presence of QRAM were investigated., Results: We found QRAM in 58 of 669 (9%) clients with an MG infection: 36 of 375 (10%) in the study population of men with urethritis and 22 of 294 (7%) in the study population of other clients (including both men and women; p=0.334). Most prevalent mutations in the parC gene were S83I and D87N, occurring in 31 of 60 (52%) and 20 of 60 (33%) samples, respectively. Factors associated with the presence of QRAM were: men who have sex with men (adjusted OR (aOR) 3.4, 95% CI 1.7 to 6.9) and Asian origin (aOR 2.5, 95% CI 1.2 to 5.6). Multidrug resistance (QRAM plus macrolide resistance-associated mutations) was found in 46 of 669 (7%) clients., Conclusions: Nine per cent of MG-positive clients from two Dutch SHCs had QRAM. New treatment strategies and antibiotics are needed to treat symptomatic patients with multidrug-resistant MG., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

Introduction: Shigellosis is a reportable infectious disease. It can present as a severe bloody diarrhoea but is often asymptomatic. Shigella can be sexually transmissible. We performed a study among symptomatic and asymptomatic men who have sex with men (MSM) to assess the prevalence of Shigella, Salmonella and Campylobacter ., Methods: From March to June 2020, MSM attending the Amsterdam centre for sexual health were consecutively included. Predefined minimal numbers of inclusion of 150, 100 and 50 were determined, respectively, for MSM who reported no diarrhoea, diarrhoea during last month or diarrhoea on the day of visit to clinic. Anal samples were tested for the presence of Shigella , Salmonella and Campylobacter . During the same period, the frequency of these bacteria was assessed in routinely tested samples requested by general physicians or nursing home physicians. Characteristics of included MSM were compared between the men with different diarrhoea anamnesis, and the prevalence of shigellosis was estimated in each group., Results: We included 212 MSM without diarrhoea, 109 MSM who recently had diarrhoea and 68 MSM who reported diarrhoea on the day of clinic visit. Thirteen (3.3%, 95% CI 1.7% to 5.6%) MSM were infected with Shigella , none with Salmonella and 7 (1.8%, 95% CI 0.7% to 3.7%) with Campylobacter. Shigella prevalence was 2.8% (95% CI 1.0% to 6.1%) in asymptomatic men, 3.7% (95% CI 1.0% to 9.1%) in men who recently had diarrhoea and 4.4% (95% CI 0.9% to 12.4%) in men with current diarrhoea (p=0.799). Shigella was more frequently found in MSM who had used pre-exposure prophylaxis (PrEP) in the preceding 3 months (10/151), compared with those not having used PrEP (2/146) or being HIV positive (1/75) (p=0.038). Shigella was significantly more often detected among MSM compared with routinely obtained faecal samples being 11/770 (1.4%) (p=0.031)., Conclusion: Shigella infections are relatively common in both symptomatic and asymptomatic MSM. Future studies should focus on the risk of onward transmission via asymptomatic persons. Samenvatting Introductie kan seksueel overdraagbaar zijn. We hebben een onderzoek uitgevoerd onder symptomatische en asymptomatische mannen die seks hebben met mannen (MSM) om de prevalentie van Shigella kan seksueel overdraagbaar zijn. We hebben een onderzoek uitgevoerd onder symptomatische en asymptomatische mannen die seks hebben met mannen (MSM) om de prevalentie van Shigella, Salmonella en Campylobacter Van maart tot juni 2020 werden achtereenvolgens MSM van het Amsterdamse centrum voor seksuele gezondheid opgenomen. Vooraf gedefinieerde minimale aantallen van inclusie van respectievelijk 150, 100 en 50 waren bepaald voor MSM die geen diarree, diarree in de afgelopen maand of diarree op de dag van bezoek aan de kliniek meldden. Anale monsters werden getest op de aanwezigheid van Methoden Van maart tot juni 2020 werden achtereenvolgens MSM van het Amsterdamse centrum voor seksuele gezondheid opgenomen. Vooraf gedefinieerde minimale aantallen van inclusie van respectievelijk 150, 100 en 50 waren bepaald voor MSM die geen diarree, diarree in de afgelopen maand of diarree op de dag van bezoek aan de kliniek meldden. Anale monsters werden getest op de aanwezigheid van Shigella, Salmonella . In dezelfde periode werd de frequentie van deze bacteriën bepaald in routinematig geteste monsters aangevraagd door huisartsen of verpleeghuisartsen. Kenmerken van geïncludeerde MSM werden vergeleken tussen mannen met verschillende diarree anamnese, en de prevalentie van shigellose werd in elke groep geschat. Campylobacter We includeerden 212 MSM zonder diarree, 109 MSM die onlangs diarree hadden en 68 MSM die diarree meldden op de dag van het bezoek aan de kliniek. Dertien (3,3%, 95% CI 1,7-5,6%) MSM waren geïnfecteerd met Resultaten We includeerden 212 MSM zonder diarree, 109 MSM die onlangs diarree hadden en 68 MSM die diarree meldden op de dag van het bezoek aan de kliniek. Dertien (3,3%, 95% CI 1,7-5,6%) MSM waren geïnfecteerd met Shigella , geen enkele met Salmonella . De prevalentie van Campylobacter . De prevalentie van Shigella was 2,8% (95%CI 1,0-6,1%) bij asymptomatische mannen, 3,7% (95%CI 1,0-9,1%) bij mannen die recent diarree hadden en 4,4% (95%CI 0,9-12,4%) bij mannen met huidige diarree (P=0,799). Shigella werd vaker gevonden bij MSM die in de voorgaande drie maanden (10/151) PrEP hadden gebruikt dan bij mensen die geen PrEP hadden gebruikt (2/146) of hiv-positief waren (1/75) (p=0,038). Shigella infecties komen relatief vaak voor bij zowel symptomatische als asymptomatische MSM. Toekomstige studies moeten zich richten op het risico van verdere overdracht via asymptomatische personen.Conclusie Shigella infecties komen relatief vaak voor bij zowel symptomatische als asymptomatische MSM. Toekomstige studies moeten zich richten op het risico van verdere overdracht via asymptomatische personen., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2022. No commercial re-use. See rights and permissions. Published by BMJ.)

Background: Syphilis diagnosis may be challenging, especially in the asymptomatic and early clinical stages. We evaluated the presence of Treponema pallidum DNA (TP-DNA) in various sample types to elucidate transmissibility during various syphilis stages., Methods: The study was conducted at the Amsterdam Centre for Sexual Health. We included adult men who have sex with men (MSM), who were suspected of having syphilis. The 2020 European guidelines definitions were followed for the diagnosis and staging of syphilis. Using a polymerase chain reaction (PCR) targeting the polA gene of Treponema pallidum (TP-PCR), we tested the following study samples on TP-DNA: peripheral blood, oropharyngeal swab, ano-rectal swab, and urine., Results: From November 2018 to December 2019 we included 293 MSM. Seventy clients had primary syphilis, 73 secondary syphilis, 86 early latent syphilis, 14 late latent syphilis, 23 treated syphilis, and 27 had no syphilis. TP-DNA was detected in at least 1 study sample in 35/70 clients with primary syphilis (2/70 peripheral blood, 7/70 oropharynx, 13/70 ano-rectum, and 24/70 urine); in 62/73 clients with secondary syphilis (15/73 peripheral blood, 47/73 oropharynx, 37/73 ano-rectum, and 26/73 urine); and in 29/86 clients with early latent syphilis (5/86 peripheral blood, 21/86 oropharynx, 11/86 ano-rectum, and 6/86 urine). TP-DNA was not detected in clients with late latent syphilis or treated syphilis, nor in clients without syphilis., Conclusions: TP-DNA was frequently detected in various sample types in the absence of lesions. This is in line with the high transmission rate of syphilis and opens diagnostic opportunities for early presymptomatic syphilis stages., Competing Interests: Potential conflicts of interest. The authors: No reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest., (© The Author(s) 2022. Published by Oxford University Press for the Infectious Diseases Society of America.)

Syphilis, caused by Treponema pallidum subspecies pallidum (TP), is a complex multistage infectious disease. Systematic dissemination occurs within a few hours of transmission. We determined the molecular variation of TP at various body locations and peripheral blood within patients in different stages of syphilis to assess the distribution of TP strains at these locations. We included 162 men who have sex with men (MSM) with syphilis visiting the Sexual Health Center in Amsterdam between 2018 to 2019, who had TP DNA detected in at least one sample type (anal swab, urine sample, peripheral blood, pharyngeal swab, and/or ulcer swab). TP DNA was detected in 287 samples using a qPCR targeting the polA gene. With multilocus sequence typing (TP-MLST) based on partial sequence analysis of three genetic regions ( tp0136, tp0548, tp0705 ), we characterized all TP DNA positive samples. Samples could be typed (119/287) from at least one anatomical location or peripheral blood from 93/162 (57%) patients in the following stages: 48 (52%) primary, 35 (38%) secondary, and 10 (11%) early latent stage syphilis. The TP-MLST type was identical within each of the 12 patients with typed samples at ≥2 different body locations. The most prevalent TP strains were 1.3.1 (39/93, 42%) and 1.1.1 (17/93, 18%) belonging to the SS14 lineage; 80% (74/93) of the patients carried a SS14 lineage TP strain and 20% (19/93) Nichols lineage. The distribution of TP-MLST types did not differ between patients by syphilis stage. We found intrapatient TP strain homogeneity and no TP strain variation between anatomical location or syphilis stages. More early latent samples should be typed and added in future studies to investigate this in more detail. IMPORTANCE Syphilis, caused by Treponema pallidum subspecies pallidum , is a complex multistage infectious disease. Systematic dissemination is known to occur within a few hours of transmission. Despite the effective antibiotic penicillin, syphilis remains prevalent worldwide. Men who have sex with men are disproportionally affected in high income countries like the Netherlands where 96% of the syphilis cases in 2020 were among this population. The inability to in vitro culture T. pallidum directly from patient samples limits whole-genome sequencing efforts. Fortunately, in 2018 a multilocus sequence typing technique was developed for T. pallidum allowing the monitoring of circulating strains. The significance of our research is in the investigation of T. pallidum molecular variation at various body locations and blood within patients in different stages of syphilis in order to assess the distribution of strains at these locations.

STD-associated questions and symptoms are submitted frequently to general practitioners and STD outpatient-clinics. In this teaching article we address 10 important clinical questions regarding epidemiology, risk assessment, testing policy, diagnostics and prevention. STD's form a separate category of infectious diseases because of the role of sexuality. Good communication about sexual behavior is indispensable for an adequate diagnosis. We discuss the recognition of extragenital manifestations of STD, which requires alertness. Estimating the STD-risk based on sexual behavior is essential for testing policy. Persons at high risk are tested for the big five. In other cases testing is based on symptoms and complaints. HIV and syphilis are serious std's. Early detection followed by treatment is important in preventing health damage and preventing further spread. Hiv-indicator-conditions are useful alarm-signs for this purpose. PrEP can help not to acquire hiv and increases sexual health. It can be prescribed by gp's and public health clinicians. But condom-use remains crucial in prevention.

BackgroundGonococcal antimicrobial resistance is emerging worldwide and is monitored in the Netherlands in 18 of 24 Sexual Health Centres (SHC).AimTo report trends, predictors and regional variation of gonococcal azithromycin resistance (AZI-R, minimum inhibitory concentration (MIC) > 1 mg/L) and ceftriaxone decreased susceptibility (CEF-DS, MIC > 0.032 mg/L) in 2013-2019.MethodsSHC reported data on individual characteristics, sexually transmitted infection diagnoses, and susceptibility testing (MIC, measured by Etest). We used multilevel logistic regression analysis to identify AZI-R/CEF-DS predictors, correcting for SHC region. Population differences' effect on regional variance of AZI-R and CEF-DS was assessed with a separate multilevel model.ResultsThe study included 13,172 isolates, predominantly (n = 9,751; 74%) from men who have sex with men (MSM). Between 2013 and 2019, annual proportions of AZI-R isolates appeared to increase from 2.8% (37/1,304) to 9.3% (210/2,264), while those of CEF-DS seemed to decrease from 7.0% (91/1,306) to 2.9% (65/2,276). Among SHC regions, 0.0‒16.9% isolates were AZI-R and 0.0-7.0% CEF-DS; population characteristics could not explain regional variance. Pharyngeal strain origin and consultation year were significantly associated with AZI-R and CEF-DS for MSM, women, and heterosexual men. Among women and heterosexual men ≥ 4 partners was associated with CEF-DS, and ≥ 10 with AZI-R.ConclusionsNo resistance or decreasing susceptibility was found for CEF, the first line gonorrhoea treatment in the Netherlands. Similar to trends worldwide, AZI-R appeared to increase. Regional differences between SHC support nationwide surveillance with regional-level reporting. The increased risk of resistance/decreased susceptibility in pharyngeal strains underlines the importance of including extragenital infections in gonococcal resistance surveillance.

Knowledge of within-host genetic variation informs studies on transmission dynamics. We studied within-host genetic variation in Neisseria gonorrhoeae over the course of infection and across different anatomical locations. Isolates were obtained during a clinical trial, and isolates from consecutive time points reflected persistent infections after treatment failure. We compared sequence types (STs) and recombination unfiltered- and filtered core genome single nucleotide polymorphism (SNP) distances in 65 within-host isolate pairs from the same anatomical location over time-obtained with a median interval of 7 days-and 65 isolate pairs across different anatomical locations at one time point. Isolates with different Multi-Locus Sequence Types (MLST), NG-Sequence Types for Antimicrobial Resistance (NG-STAR) and NG-Multi Antigen Sequence Types (NG-MAST) had a median of 1466 recombination filtered SNPs, whereas a median of 1 SNP was found between isolates with identical STs or a different NG-MAST only. The threshold for differentiating between strains was set at 10 recombination filtered SNPs, showing that isolates from persistent infections could have different NG-MASTs. Antibiotic pressure applied through treatment did not lead to an increase in genetic variation in specific genes or in overall extent of variation, compared to variation across anatomical locations. Instead, within-host genetic variation was proposedly driven by the host immune response, as it was concentrated in genomic regions encoding surface exposed proteins involved in host-microbe interaction. Ultimately, 15/228 (6.5%) between-host pairs contained a single strain, suggesting between-host transmission. However, patient reported data are needed to differentiate within-host persistence from between-host transmission. IMPORTANCE Understanding transmission dynamics of Neisseria gonorrhoeae ( Ng ) is based on the identification of transmission events. These can be identified by assessing genetic relatedness between Ng isolates, expressed as core genome SNP distances. However, a SNP threshold to differentiate between strains needs to be defined, using knowledge on within- and between-host genetic variation. Here, we assessed within-host genetic variation, using a unique set of within-host Ng isolates from the same anatomical location over time or across different anatomical locations at one time point. The insights in genetic variation that occurred during the infection period contribute to the understanding of infection dynamics. In addition, the obtained knowledge can be used for future research on transmission dynamics and development of public health interventions based on bacterial genomic data.

The bacterial accessory genome provides the genetic flexibility needed to facilitate environment and host adaptation. In Neisseria gonorrhoeae, known accessory elements include plasmids which can transfer and mediate antimicrobial resistance (AMR); however, chromosomal accessory genes could also play a role in AMR. Here, the gonococcal accessory genome was characterized using gene-by-gene approaches and its association with the core genome and AMR were assessed. The gonococcal accessory gene pool consisted of 247 genes, which were mainly genes located on large mobile genetic elements, phage associated genes, or genes encoding putative secretion systems. Accessory elements showed similar synteny across genomes, indicating either a predisposition for particular genomic locations or ancestral inheritance that are conserved during strain expansion. Significant associations were found between the prevalence of accessory elements and core genome multi-locus sequence types (cgMLST), consistent with a structured gonococcal population despite frequent horizontal gene transfer (HGT). Increased prevalence of putative DNA exchange regulators was significantly associated with AMR, which included a putative secretion system, methyltransferases and a toxin-antitoxin system. Although frequent HGT results in high genetic diversity in the gonococcus, we found that this is mediated by a small gene pool. In fact, a highly organized genome composition was identified with a strong association between the accessory and core genome. Increased prevalence of DNA exchange regulators in antimicrobial resistant isolates suggests that genetic material exchange plays a role in the development or maintenance of AMR. These findings enhance our understanding of gonococcal genome architecture and have important implications for gonococcal population biology. IMPORTANCE The emergence of antimicrobial resistance (AMR) against third generation cephalosporins in Neisseria gonorrhoeae is a major public health concern, as these are antibiotics of last resort for the effective treatment of gonorrhea. Although the resistance mechanisms against this class of antibiotics have not been entirely resolved, resistance against other classes of antibiotics, such as tetracyclines, is known to be mediated through plasmids, which are known gonococcal extra-chromosomal accessory elements. A complete assessment of the chromosomal accessory genome content and its role in AMR has not yet been undertaken. Here, we comprehensively characterize the gonococcal accessory genome to better understand genome architecture as well as the evolution and mechanisms of AMR in this species.

Background: Mycoplasma genitalium (MG) is associated with urethritis in men and could play a role in clinical outcome. We examined clinical improvement of symptoms in men receiving empirical treatment for urethritis and correlated the outcome with Neisseria gonorrhoeae (NG), Chlamydia trachomatis (CT), MG, and MG macrolide resistance-associated mutations (MRAM) status., Methods: At the sexually transmitted infection clinic in Amsterdam, the Netherlands, empirical treatment for gonococcal urethritis is 1 g ceftriaxone and for nongonococcal urethritis 1 g azithromycin. In 2018 to 2019, we tested urine samples of men with urethritis for CT, NG, and MG using transcription-mediated amplification assays. Mycoplasma genitalium-positive samples were tested for MRAM using quantitative polymerase chain reaction. Two weeks after receiving therapy, men were sent a text message inquiring after clinical improvement., Results: We evaluated 2505 cases of urethritis. The positivity rates of NG, CT, and MG were 26% (648 of 2489), 29% (726 of 2489), and 23% (522 of 2288), respectively. In 768 of 2288 of the cases (34%), no causative agent was detected. Most cases were infected with a single pathogen: NG, 417 of 2288 (18%); CT, 486 of 2288 (21%); and MG, 320 of 2288 (14%). The prevalence of MRAM among MG-positives was 74% (327 of 439). For 642 (25.6%) cases, we could evaluate clinical improvement after treatment of whom 127 (20%) indicated no improvement; 9% (15 of 174) in NG cases, 18% (35 of 195) in CT cases, 14% (4 of 28) in MG wild-type cases, and 40% (38 of 94) in MG-MRAM cases. Clinical improvement in MG-MRAM cases was significantly lower compared with all other groups (P < 0.001)., Conclusions: Presence of MG-MRAM is associated with lack of clinical improvement in azithromycin-treated nongonococcal urethritis., Competing Interests: Conflict of Interest and Sources of Funding: None declared., (Copyright © 2021 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Sexually Transmitted Diseases Association.)

Background: Neisseria gonorrhoeae causes gonorrhoea, a common sexually transmitted infection. Emerging strains resistant to first-line ceftriaxone threaten N gonorrhoeae management. Hence, alternative treatments are needed. We aimed to evaluate the efficacy of ertapenem, gentamicin, and fosfomycin as alternative treatments for anogenital N gonorrhoeae., Methods: In a randomised, controlled, double-blind, non-inferiority trial (three experimental groups and one control group) at the Centre for Sexual Health in Amsterdam, Netherlands, we included adults aged 18 years or older, with anorectal or urogenital gonorrhoea. With random permuted blocks, participants were randomly assigned (1:1:1:1) to receive intramuscular 500 mg ceftriaxone (control group), intramuscular 1000 mg ertapenem, intramuscular 5 mg/kg gentamicin (maximum 400 mg), or oral 6 g fosfomycin. The primary outcome was the proportion of participants with a negative nucleic acid amplification test of the predefined primary infected site, 7-14 days after treatment. The primary analysis was per protocol (ie, excluding those lost to follow-up). The modified intention-to-treat analysis included all randomly assigned patients with anogenital gonorrhoea considering those lost-to-follow-up as treatment failure. Non-inferiority was established if the lower Hochberg-corrected 95% CI for difference between the experimental and control groups was greater than -10%. For the analysis of adverse events, we included all participants who received medication. The trial was registered at ClinicalTrials.gov (NCT03294395) and is complete., Findings: Between Sept 18, 2017, and June 5, 2020, from 2160 patients invited to participate, we assigned 346 (16%) participants to receive either ceftriaxone (n=103), ertapenem (n=103), gentamicin (n=102), or fosfomycin (n=38). The fosfomycin group was terminated early after interim analysis revealed less than 60% efficacy. In the primary per-protocol analysis, 93 (100%) of 93 patients in the ceftriaxone group, 86 (99%) of 87 patients in the ertapenem group, 79 (93%) of 85 patients in the gentamicin group, and four (12%) of 33 patients in the fosfomycin group cleared N gonorrhoeae (risk difference vs ceftriaxone -0·01 [95% CI -0·08 to 0·05] for ertapenem and -0·07 [-0·16 to -0·01] for gentamicin). Thus, ertapenem proved non-inferior to ceftriaxone. In mITT analysis, risk differences versus ceftriaxone were -0·08 (-0·17 to 0·003) for ertapenem and -0·11 (-0·21 to -0·04) for gentamicin. We observed a higher proportion of patients with at least one adverse event in the ertapenem group (58 [56%] of 103) and fosfomycin group (36 [95%] of 38) versus the ceftriaxone group (24 [23%] of 103)., Interpretation: Single-dose 1000 mg ertapenem is non-inferior to single-dose 500 mg ceftriaxone in gonorrhoea treatment. Yet, 5 mg/kg gentamicin (maximum 400 mg) is not non-inferior to ceftriaxone. Ertapenem is a potential effective alternative for anogenital N gonorrhoeae infections and merits evaluation for ceftriaxone-resistant infections., Funding: ZonMw and GGD-Amsterdam., Translation: For the Dutch translation of the abstract see Supplementary Materials section., Competing Interests: Declaration of interests VWJ, TH, CMW, E Hoornenborg, JdK-E, M KnolMdL, R A A Mathôt, S Bruisten, and APvD had nothing to disclose. AB reports grants from ANRS, grants from SIDACTION, outside the submitted work. MFSvdL reports grants from ZonMw, during the conduct of the study. HJCdV reports grants from ZonMw, grants from Public Health Services Amsterdam, during the conduct of the study., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

In this retrospective study, the performance of nine serological screening assays for Lyme borreliosis (LB) diagnostics was evaluated using a study population of LB cases and controls. Sera derived from 74 well-defined LB cases and 122 controls were included. The LB cases were diagnosed with erythema migrans (EM; n  = 11), Lyme neuroborreliosis (LNB; n  = 35), Lyme arthritis (LA; n  = 20), or acrodermatitis chronica atrophicans (ACA; n  = 8). Controls comprised 74 age- and gender-matched healthy individuals and 48 patients with other diseases with anticipated high rates of cross-reactivity. The assays under evaluation were selected based on a literature review and expected continued availability with CE marking under the new in vitro diagnostic regulation (European Union) 2017/746. The overall sensitivity (IgG and IgM results combined) among LB cases ranged between 54.5% (6 of 11) and 90.9% (10 of 11) for EM patients and between 97.1% (34 of 35) and 100% for patients with LNB, LA, and ACA. The positivity rate ranged between 8.1% (6 of 74) and 29.7% (22 of 74) among the healthy controls and between 22.9% (11 of 48) and 64.6% (31 of 48) among the cross-reactivity controls. The IgM results were more heterogeneous than the IgG and IgM/IgG results and did not contribute to the overall sensitivity but substantially increased the positivity rates among the controls. In conclusion, all evaluated Borrelia serological screening assays performed comparably with respect to early- and late-disseminated LB. The addition of an IgM assay to the screening of Borrelia -specific IgG antibodies had no added value for the diagnosis of Lyme borreliosis. IMPORTANCE Serology plays an important role in the diagnosis of Lyme borreliosis. Guidelines prescribe a two-tier testing algorithm in which a highly sensitive screening assay is used for screening and reactive sera are retested with an immunoblot to reduce false positivity rates. Recently, two commonly used screening assays were discontinued, including the very well-performing C6 Lyme enzyme-linked immunosorbent assay (ELISA) (Immunetics). This study provides an evaluation of the performance of nine different Borrelia serology screening assays, eight with expected future availably and the C6 Lyme ELISA, using a well-defined study panel of Lyme borreliosis patients, healthy population controls, and cross-reactivity controls. Evaluation data on multiple assays aid diagnostic laboratories in their choice for a reliable Borrelia serology screening assay to improve their diagnostic algorithm for Lyme borreliosis.

Competing Interests: We declare no competing interests.

This retrospective case-control study assesses the sensitivity, specificity, and area under the curve of the ZEUS Borrelia VlsE1/pepC10 assay in comparison with the C6-ELISA in European patients with Lyme borreliosis, healthy blood donors, and potentially cross-reactive controls. We included a convenience series of 161 sera from patients with physician-confirmed early localized or disseminated Lyme borreliosis (n = 143), 400 sera from healthy blood donors and 44 sera with potentially cross-reactive antibodies, on which we performed the aforementioned serological assays and the recomLine immunoblot. Diagnostic parameters were compared in various single-tier and two-tier algorithms. The specificities of the C6-ELISA and the ZEUS Borrelia VlsE1/pepC10 were comparable in healthy blood donors (e.g., single-tier permissive: C6: 362/400, 90.5% [87.2-93.2]; VlsE1/pepC10: 361/400, 90.3% [86.9-93.0]). The C6-ELISA had an apparently higher sensitivity in EM sera (e.g., both time points combined: C6: 61/76, 80.3% [69.5-88.5]; VlsE1/pepC10: 54/76, 71.1% [59.5-80.9]), but these differences were all not-significant. Interestingly, the VlsE1/pepC10 assay had a significantly higher specificity in sera with potentially cross-reactive antibodies (e.g., single-tier permissive: C6: 34/44, 77.3% [62.2-88.5]; VlsE1/pepC10: 40/44, 90.9% [78.3-97.5]; p = 0.031). While the areas under the curve for both assays were excellent, that of the C6-ELISA exceeded that of the VlsE1/pepC10 (C6: AUC = 0.925; VlsE1/pepC10: AUC = 0.878; p = 0.003). The novel ZEUS Borrelia VlsE1/pepC10 assay has generally comparable diagnostic parameters to the C6-ELISA with potentially improved specificity in cross-reactive sera. Thus, it is a useful tool for the serodiagnosis of Lyme borreliosis in Europe., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Objectives: To assess the in vitro effect of select antimicrobials on the growth of N. gonorrhoeae and its pharmacodynamic parameters., Methods: Time-kill assays were performed on two reference N. gonorrhoeae strains (ceftriaxone-resistant WHO X and ceftriaxone-susceptible WHO F) and one clinical N. gonorrhoeae strain (ceftriaxone-susceptible CS03307). Time-kill curves were constructed for each strain by measuring bacterial growth rates at doubling antimicrobial concentrations of ceftriaxone, ertapenem, fosfomycin and gentamicin. Inputs from these curves were used to estimate minimal bacterial growth rates at high antimicrobial concentrations ( ψ min ), maximum bacterial growth rates in the absence of antimicrobials ( ψ max ), pharmacodynamic minimum inhibitory concentrations (zMIC), and Hill's coefficients (κ)., Results: Ceftriaxone, ertapenem and fosfomycin showed gradual death overtime at higher antimicrobial concentrations with a relatively high ψ min , demonstrating time-dependent activity. Compared to WHO F, the ψ min for WHO X was significantly increased, reflecting decreased killing activity for ceftriaxone, ertapenem and fosfomycin. At high ceftriaxone concentrations, WHO X was still efficiently killed. CS03307 also showed a high ψ min for ceftriaxone in spite of a low MIC and no difference in ψ min for fosfomycin in spite of significant MIC and zMIC differences. Gentamicin showed rapid killing for all three strains at high concentrations, demonstrating concentration-dependent activity., Conclusions: Based on time-kill assays, high-dosage ceftriaxone could be used to treat N. gonorrhoeae strains with MIC above breakpoint, with gentamicin as a potential alternative. Whether ertapenem or fosfomycin would be effective to treat strains with a high MIC to ceftriaxone is questionable.

Competing Interests: MEB and JH: LB diagnostics in collaboration with various companies, although none of that work involved molecular detection of B. burgdorferi sensu lato. MEB and JH have not received any personal compensation from any of said companies, nor were any of said companies involved in any aspect of the current manuscript. VF: Research support: RKI/BMG, ESCMID, ECDC, StMGP/StMUG, INSTAND. Lecturing activities (honoraria, travel expenses): DIAMEDIS, Diasorin, Mikrogen, Seramun, Siemens, HLR. Consulting activities EQA schemes (no honoraria): QCMD, INSTAND, ECDC. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Background: It is important to gain insight into the burden of COVID-19 at city district level to develop targeted prevention strategies. We examined COVID-19 related hospitalisations by city district and migration background in the municipality of Amsterdam, the Netherlands., Methods: We used surveillance data on all PCR-confirmed SARS-CoV-2 hospitalisations in Amsterdam until 31 May 2020, matched to municipal registration data on migration background. We calculated directly standardised (age, sex) rates (DSR) of hospitalisations, as a proxy of COVID-19 burden, per 100,000 population by city district and migration background. We calculated standardised rate differences (RD) and rate ratios (RR) to compare hospitalisations between city districts of varying socio-economic and health status and between migration backgrounds. We evaluated the effects of city district and migration background on hospitalisation after adjusting for age and sex using Poisson regression., Results: Between 29 February and 31 May 2020, 2326 cases (median age 57 years [IQR = 37-74]) were notified in Amsterdam, of which 596 (25.6%) hospitalisations and 287 (12.3%) deaths. 526/596 (88.2%) hospitalisations could be matched to the registration database. DSR were higher in individuals living in peripheral (South-East/New-West/North) city districts with lower economic and health status, compared to central districts (Centre/West/South/East) (RD = 36.87,95%CI = 25.79-47.96;RR = 1.82,95%CI = 1.65-1.99), and among individuals with a non-Western migration background compared to ethnic-Dutch individuals (RD = 57.05,95%CI = 43.34-70.75; RR = 2.36,95%CI = 2.17-2.54). City district and migration background were independently associated with hospitalisation., Conclusion: City districts with lower economic and health status and those with a non-Western migration background had the highest burden of COVID-19 during the first wave of COVID-19 in Amsterdam., (© 2021. The Author(s).)

Mycoplasma genitalium is a well-known cause of urethritis in men and has been associated with cervicitis, pelvic inflammatory disease, and adverse obstetric outcomes in women. In this cross-sectional study, we determined the current prevalence of M. genitalium infection and the rate of macrolide resistance in M. genitalium isolates, in patients visiting two large Dutch sexually transmitted infection (STI) clinics, to evaluate whether the recommendations in Dutch guidelines should be revised. In addition, risk factors for M. genitalium were identified. In total, 3225 patients were included. M. genitalium prevalence rates were 13.8% for all patients; 20.1% for men who have sex with men, 8.2% for men who have sex with women, and 12.6% for women. Macrolide resistance-associated mutations were detected in 66% of the patients infected with M. genitalium . Age, educational level, country of origin, number of sexual partners, HIV-positivity, infection with Neisseria gonorrhoeae , and urethral symptoms in men were independently associated with M. genitalium infection. In conclusion, we found very high prevalence rates and macrolide resistance rates of M. genitalium in patients visiting STI clinics.

Background: Emerging resistance to cephalosporins in Neisseria gonorrhoeae (Ng) is a major public health threat, since these are considered antibiotics of last resort. Continuous surveillance is needed to monitor the circulation of resistant strains and those with reduced susceptibility., Objectives: For the purpose of epidemiological surveillance, genomic population analysis was performed on Ng isolates from Amsterdam with a focus on isolates with reduced susceptibility to ceftriaxone., Methods: WGS data were obtained from 318 isolates from Amsterdam, the Netherlands between 2014 and 2019. Isolates were typed according to MLST, Ng Multi-Antigen Sequence Typing (NG-MAST) and Ng Sequence Typing for Antimicrobial Resistance (NG-STAR) schemes and additional resistance markers were identified. Phylogenetic trees were created to identify genetic clusters and to compare Dutch and non-Dutch MLST7827 isolates., Results: MLST7363 and MLST1901 were the predominant strains having reduced susceptibility to ceftriaxone during 2014-16; MLST7827 emerged and dominated during 2017-19. NG-STAR38 and NG-MAST2318/10386 were predominant among MLST7827 isolates. MLST7827 reduced susceptibility isolates carried a non-mosaic 13.001 penA allele with an A501V mutation and porB1b G120K/A121D mutations, which were lacking in susceptible MLST7827 isolates. Phylogenetic analysis of all publicly available MLST7827 isolates showed strong genetic clustering of Dutch and other European MLST7827 isolates., Conclusions: MLST7827 isolates with reduced ceftriaxone susceptibility have emerged during recent years in Amsterdam. Co-occurrence of penA A501V and porB1b G120K/A121D mutations was strongly associated with reduced susceptibility to ceftriaxone. Genetic clustering of Dutch and other European MLST7827 isolates indicates extensive circulation of this strain in Europe. Close monitoring of the spread of this strain having an alarming susceptibility profile is needed., (© The Author(s) 2021. Published by Oxford University Press on behalf of the British Society for Antimicrobial Chemotherapy.)

Background: Extended-spectrum β-lactamase Enterobacteriaceae (ESBL-E) may be sexually transmitted. Men who have sex with men (MSM) have different sexual behaviour than the general population, and thus may be at risk for ESBL-E carriage. This study determined the prevalence of ESBL-E carriage and its association with sexual behaviour among MSM in Amsterdam, The Netherlands., Materials and Methods: In total, 583 HIV-positive and HIV-negative MSM from the Amsterdam Cohort Study were screened for rectal ESBL-E carriage between April and December 2018. Participants completed a self-administered questionnaire on (sexual) behaviour and risk factors for antimicrobial resistance. The proportion of the study population with ESBL-E carriage was compared by number of sexual partners using logistic regression, and across clusters of sexual behaviours with steady and casual partners, separately, using latent class analyses; all results were adjusted for recent use of antibiotics, travel and hospitalization., Results: Overall, 16.3% [95% confidence interval (CI) 13.4-19.5] of the study population tested positive for ESBL-E. The odds of ESBL-E carriage increased as number of sexual partners increased [adjusted odds ratio per ln(partner+1), 1.57, 95% CI 1.26-1.94; P<0.001]. There was no association between ESBL-E carriage and sexual behaviour with steady partner(s). Compared with participants in the 'no sex with casual partner(s)' cluster, adjusted odds of being ESBL-E positive were 2.95-fold higher (95% CI 1.52-5.80) for participants in the 'rimming and frottage' cluster (P=0.001) and 2.28-fold higher (95% CI 0.98-5.31) for participants in the 'toy use and fisting' cluster (P=0.056)., Conclusions: The prevalence of ESBL-E in MSM is higher compared with the overall Dutch population, likely due to sexual transmission with casual partners. This implies that sexually active MSM should be considered a risk group for ESBL-E carriage., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Background: Surveillance of recent HIV infections (RHI) using an avidity assay has been implemented at Dutch sexual health centres (SHC) since 2014, but data on RHI diagnosed at other test locations is lacking. Setting: Implementation of the avidity assay in HIV treatment clinics for the purpose of studying RHI among HIV patients tested at different test locations. Methods: We retrospectively tested leftover specimens from newly diagnosed HIV patients in care in 2013-2015 in Amsterdam. Avidity Index (AI) values ≤0.80 indicated recent infection (acquired ≤6 months prior to diagnosis), and AI > 0.80 indicated established infection (acquired >6 months prior to diagnosis). An algorithm for RHI was applied to correct for false recency. Recency based on this algorithm was compared with recency based on epidemiological data only. Multivariable logistic regression analysis was used to identify factors associated with RHI among men who have sex with men (MSM). Results: We tested 447 specimens with avidity; 72% from MSM. Proportions of RHI were 20% among MSM and 10% among heterosexuals. SHC showed highest proportions of RHI (27%), followed by GPs (15%), hospitals (5%), and other/unknown locations (11%) ( p < 0.001). Test location was the only factor associated with RHI among MSM. A higher proportion of RHI was found based on epidemiological data compared to avidity testing combined with the RHI algorithm. Conclusion: SHC identify more RHI infections compared to other test locations, as they serve high-risk populations and offer frequent HIV testing. Using avidity-testing for surveillance purposes may help targeting prevention programs, but the assay lacks robustness and its added value may decline with improved, repeat HIV testing and data collection., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Slurink, van de Baan, van Sighem, van Dam, van de Laar, de Bree, van Benthem and Op de Coul.)

Mycoplasma genitalium (MG) is a sexually transmitted bacterium in which macrolide resistance is rapidly increasing, limiting treatment options. We validated a new assay to detect the presence of macrolide resistance-associated mutations in MG (MG-MRAM). In 2018, symptomatic and asymptomatic clients visiting sexually transmitted infections (STI) clinics in Amsterdam or The Hague were tested for MG using transcription mediated amplification (TMA) assays. The sensitivity to detect MG of the newly developed MG-MRAM qPCR was compared to the MgPa qPCR, both in relation to the TMA assay. For the sensitivity and specificity to detect relevant mutations the MG-MRAM qPCR was compared to 23SrRNA sequencing analysis. The qPCR was subsequently used to determine the presence of MG-MRAM at different anatomical locations and to identify risk factors for MG-MRAM. MG-positive clients (402) providing 493 MG-positive samples were included. In total 309/493 (62.7%) samples from 291 (72.4%) clients were successfully typed with the MG-MRAM qPCR. The MG-MRAM qPCR had a sensitivity of 98.6% (95%CI 91.1%-99.9%) and specificity of 94.1% (95%CI 78.9%-99.0%) to detect MG-MRAM compared to sequencing analysis. Infection with MG-MRAM was detected in 193/291 (66.3%) clients: in 129/178 (72.5%) men and 64/113 (56.6%) women (p = 0.005). Prevalence of MG-MRAM was significantly higher in men, clients with a higher education, HIV-positive clients and clients with >10 sexual partners in the previous six months, but in multivariable analysis no factor was significantly associated with MG-MRAM presence. Since MG-MRAM prevalence was very high, testing for MG-MRAM is essential if treatment for MG is considered, and can be performed with this sensitive and specific qPCR test in routine diagnostics., Competing Interests: The authors have read the journal’s policy and have the following potential competing interests: Hologic Inc, San Diego, USA provided part of the diagnostic tests to detect MG. This does not alter our adherence to PLOS ONE policies on sharing data and materials. There are no patents, products in development or marketed products associated with this research to declare.

Background: Nasal carriage of methicillin-resistant Staphylococcus aureus (MRSA) is associated with an increased risk of infection. Colonization with MRSA is observed in < 1% of the general Dutch population. Increased risk for MRSA carriage is known to occur in several key groups, one of which is asylum seekers. However, little is known about MRSA carriage among undocumented migrants and uninsured legal residents. This study aimed to determine the prevalence of nasal MRSA carriage among these groups in Amsterdam, the Netherlands., Methods: In this cross-sectional study, between October 2018 and October 2019, undocumented migrants and uninsured legal residents aged 18 years or older who were able to understand one of the study languages were recruited at an NGO health care facility in Amsterdam, the Netherlands, for general practitioner (GP) consultations. Participants were asked questions on demographics, migration history, antibiotic use and other possible risk factors for MRSA carriage and were screened for nasal MRSA carriage by selective culturing e-swabs. Characteristics of MRSA-negative and MRSA-positive participants were compared using univariable logistic regression analysis with Firth's correction., Results: Of the 3822 eligible patients, 760 were screened for nasal MRSA carriage (19.9%). Of the 760 participants, over half were male (58%; 442/760) and originated mainly from Africa (35%; 267/760), Asia (30%; 229/760) and North or South America (30%; 227/760). In total, 705/760 participants (93%) were undocumented migrants and 55/760 (7%) were uninsured legal residents of Amsterdam. The overall prevalence of nasal MRSA carriage was 2.0% (15/760) (95%CI 1.1 to 3.2%), with no difference between undocumented migrants (14/705) (2.0, 95%CI 1.1 to 3.3%) and uninsured legal residents (1/55) (1.8, 95%CI 0.1 to 9.7%). Genotyping showed no clustering of the 15 isolates. MRSA carriage was not associated with sociodemographic, migration history or other possible risk factors. Nevertheless, this study had limited power to detect significant determinants. Three participants (3/15; 20%) harbored Panton-Valentine leukocidin (PVL)-positive isolates., Conclusion: Even though our study population of undocumented migrants and uninsured legal residents had a higher prevalence of nasal MRSA carriage compared to the general Dutch population, the prevalence was relatively low compared to acknowledged other high-risk groups.

Background: Syphilis is a re-emerging infection. Sexually transmitted infection (STI) clinics and GPs are important providers of STI care in the Netherlands. The role of GPs in syphilis care is assumed to be small, since most men who have sex with men (MSM) visit STI clinics for STI care., Aim: To explore the role of GPs in the diagnosis and treatment of syphilis., Design & Setting: Data on syphilis diagnostics by GPs in Amsterdam between 2011 and 2017 were retrieved from laboratories, covering 90% of the GPs. The study also used the academic GPs' network database to explore the management of syphilis by GPs between 2013 and 2018., Method: Syphilis tests requested by GPs were analysed and compared with annual reports of the STI clinic. Patients with an International Classification of Primary Care-1 (ICPC-1) syphilis code were identified in the GP database. Cases diagnosed by the GP were evaluated whether they were treated by the GP or referred to secondary care., Results: In the laboratory database, GPs had diagnosed syphilis 522 times, compared with 2515 times by the STI clinics. Based on the 90% coverage of GPs, the contribution of all Amsterdam's GPs was 19% of the total number of diagnoses. Consequently, the annual incidence of syphilis diagnosed by the GP was 10.2 per 100 000 inhabitants. Of the 43 cases identified in the GP database, six (14.0%) were referred and 33 (76.7%) were treated by a GP., Conclusion: Although for an individual GP, syphilis is rare to diagnose, GPs in Amsterdam do contribute to the rate of syphilis diagnosis and appear to treat the majority of cases that they have diagnosed., (Copyright © 2020, The Authors.)

Objectives: The prevalence of syphilis, caused by the spirochete Treponema pallidum subsp . pallidum (TPA), remains high despite the availability of effective antibiotics. In the Netherlands, most syphilis cases are found among men who have sex with men (MSM). We studied the distribution of TPA strain types by molecular characterisation and related this to available characteristics. In addition, resistance to macrolides was assessed., Methods: TPA DNA was extracted from 136 genital ulcer swab or skin lesions samples deriving from 135 patients diagnosed with syphilis in 2016 and 2017 at the Public Health Service in Amsterdam, the Netherlands. Molecular typing was done according to the enhanced CDC method (E-CDC), in which three genetic regions of the arp , tpr and tp0548 genes are analysed by gel electrophoresis of the arp and tpr regions and by sequence analysis for the tp0548 region. Part of the 23S rDNA locus was sequenced to determine the presence of macrolide resistance-associated mutations., Results: Full E-CDC strain types could be determined for 99/136 (73%) DNA samples, which tested positive in a diagnostic PCR targeting the polA gene. Types differed within one patient of whom two samples were available. No association was found between the demographic and clinical characteristics and the TPA types. The most prevalent type was 14d/g, found in 23 of the 99 (23%) fully typed samples. Part of the 23S rDNA locus was successfully sequenced for 93/136 (68%) samples and 83 (88%) contained the A2058G mutation. No A2059G mutation was found., Conclusions: A broad strain distribution was found. Few subtypes were clonally expanded, and most other subtypes were rare. Detection of the most prevalent strain type, 14d/g, is in concordance with other TPA typing studies. The high prevalence of genetic macrolide resistance indicates that azithromycin is not an alternative treatment option., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY. Published by BMJ.)

Background: Absence of rapid antimicrobial resistance testing of Neisseria gonorrhoeae (Ng) hinders personalized antibiotic treatment. To enable rapid ciprofloxacin prescription, a real-time polymerase chain reaction (PCR) for simultaneous detection of Ng and fluoroquinolone resistance-associated gyrA-S91F mutation was evaluated., Methods: Analytical NG quantitative PCR kit (NYtor BV) performance was assessed on 50 Ng transcription-mediated amplification (TMA)-negative and 100 Ng TMA-positive samples. To assess clinical use, 200 samples were prospectively analyzed, in parallel to routine diagnostic tests. Also, 50 urine, 50 anal, 50 pharyngeal, and 50 vaginal Ng TMA-positive samples were retrospectively analyzed. To assess if patients carried strains with different ciprofloxacin sensitivity at different anatomical locations, 50 urine/anal or vaginal/anal sample pairs collected during a single visit were analyzed., Results: The NG quantitative PCR kit showed 97% sensitivity and 100% specificity for Ng detection and 92% sensitivity and 99% specificity for gyrA-S91F detection. Relative to TMA results, 85% Ng detection sensitivity and 99% specificity were found. Regarding the 200 prospectively analyzed clinical samples, 13 were Ng positive, of which 10 were also tested for antibiotic susceptibility by culture. The kit showed concordance for GyrA-S91F detection in 9 of 10 samples. Ng was detected in 96% and 94% of vaginal and urine TMA-positive samples, in 84% of anal samples and only in 22% of pharyngeal samples. Discordant ciprofloxacin sensitivity was found for 2 of 26 characterized urine/anal sample pairs., Conclusion: The NG quantitative polymerase chain reaction (qPCR) kit can be implemented in diagnostic testing for vaginal, urine, and anal Ng TMA-positive samples to enable rapid prescription of oral ciprofloxacin.

Objectives: To compare molecular and epidemiological differences between ceftriaxone-reduced susceptible (CRO-RS) and ceftriaxone-susceptible (CRO-S) N. gonorrhoeae (Ng) and to study the genetic relatedness of CRO-RS isolates., Methods: Demographic and clinical data and samples for cultures were routinely collected from gonorrhoea patients visiting the Amsterdam STI clinic in 2009 to 2017. Ng multiantigen sequence typing (NG-MAST) and penA types were compared between CRO-RS and CRO-S Ng (frequency matched on year of isolation and sexual risk group). Minimum spanning trees were produced based on multilocus variable number of tandem repeats analysis for Ng (NG-MLVA) genotypes., Results: We selected 174 CRO-RS isolates (minimum inhibitory concentration, ≥0.064 mg/L) and 174 CRO-S isolates (minimum inhibitory concentration, ≤0.016 mg/L). Demographic and clinical characteristics of patients were overall comparable between those infected with CRO-RS Ng and CRO-S Ng. However, CRO-RS isolates were more often collected from the pharyngeal site (odds ratios [OR], 3.64; P < 0.001), and patients with CRO-RS Ng were less often human immunodeficiency virus (HIV) and syphilis positive (OR, 0.63; P = 0.041 and OR, 0.58; P = 0.028, respectively). We identified 12 clusters based on NG-MLVA genotypes, including 3 large (>25 isolates) clusters predominantly containing CRO-RS isolates. Those from cluster 1 (n = 32) were mostly from 2009 to 2012 (n = 24; 75.0%), with a mosaic penA XXXIV pattern (n = 27; 84.4%) and belonging to NG-MAST genogroup G1407 (n = 24; 75.0%). Isolates from cluster 2 (n = 29) were mostly from 2013 to 2015 (n = 24; 82.7%), had a nonmosaic penA IX + A501T mutation (n = 22; 75.9%) and NG-MAST G2400 (n = 14; 48.3%). Most isolates from cluster 3 (n = 37) were from 2015 to 2017 (n = 26; 70.2%), had a nonmosaic penA IV + A501V mutation (n = 24; 64.9%) and NG-MAST G2318 (n = 22; 59.5%)., Conclusions: We observed a shift in the predominant penA (from mosaic toward nonmosaic plus A501T/V mutation), NG-MAST and NG-MLVA types among CRO-RS Ng over time. This indicates a successive spread of different CRO-RS Ng clones.

Introduction: Pharyngeal Neisseria gonorrhoeae infections are mostly asymptomatic, yet sustain ongoing gonococcal transmission. We assessed the proportion of pharyngeal gonorrhea that spontaneously clears and identified determinants of clearance., Methods: At the sexually transmitted infections clinic Amsterdam, at-risk women and men who have sex with men were routinely screened for pharyngeal N. gonorrhoeae using an RNA-based nucleic acid amplification test (NAAT; Aptima Combo 2).We retrospectively examined medical records of pharyngeal gonorrhea patients (January 2012-August 2015). We included patients who returned for antibiotic treatment and had a new sample taken for NAAT before treatment. Spontaneous clearance was defined as a negative NAAT result at the follow-up visit., Results: During the study period, 1266 cases with a pharyngeal gonorrhea were not treated at the first consultation and returned for a follow-up visit. Median (interquartile range) time between the first consultation and follow-up was 10 (7-14) days. Spontaneous clearance was found in 139 cases (11.0%) and was associated with age at least 45 years (vs. 16-24 years; adjusted odds ratio, 2.02 [95% confidence interval, 1.09-3.75]) and with time from the first consultation to follow-up (adjusted odds ratio, 1.08 [1.06-1.10], per extra day)., Conclusions: Eleven percent of pharyngeal gonorrhea cases cleared spontaneously. Spontaneous clearance of pharyngeal gonorrhea was more often seen among older patients.

Background Neisseria gonorrhoeae antibiotic resistance surveillance is important to maintain adequate treatment. We analysed 2007-15 data from the Gonococcal Resistance to Antimicrobials Surveillance (GRAS), which currently includes 19 of 25 sexually transmitted infection (STI) centres in the Netherlands. Methods : From each patient with a gonorrhoea culture, the minimum inhibitory concentration (MIC) for several antibiotics was determined. Time trends were assessed by geometric means and linear regression of logarithmic MIC. Determinants for decreased susceptibility to ceftriaxone (MIC > 0.032 mg/L) and resistance to cefotaxime (MIC > 0.125 mg/L) and azithromycin (MIC > 0.5 mg/L) were assessed using stratified logistic regression. Results : 11,768 isolates were analysed. No ceftriaxone resistance was found. In 2015, 27 of 1,425 isolates (1.9%) were resistant to cefotaxime and 176 of 1,623 (10.9%) to azithromycin. Ceftriaxone susceptibility showed no trend (p = 0.96) during the study period, but cefotaxime MIC decreased (p < 0.0001) and azithromycin MIC increased (p < 0.0001) significantly. Concerning ceftriaxone, isolates of men who have sex with men (MSM) from 2013 (p = 0.0005) and 2014 (p = 0.0004) were significantly associated with decreased susceptibility. Significant determinants for cefotaxime resistance were having ≥ 6 partners for women (p = 0.0006). For azithromycin, isolates from MSM collected in 2012 (p = 0.0035), 2013 (p = 0.012), and 2014 (p = 0.013), or from non-Dutch (p < 0.0001) or older (≥ 35 years; p = 0.01) MSM were significantly associated with susceptibility. Resistance in heterosexual men was significantly associated with being ≥ 25 years-old (p = 0.0049) or having 3-5 partners (p = 0.01). Conclusions: No ceftriaxone resistance was found, but azithromycin MIC increased in 2007-15. Resistance determinants could help with focused intervention strategies.

Background: One third of travellers to low- and middle-income regions of the tropics and subtropics become colonized by extended-spectrum beta-lactamase-producing Enterobacteriaceae (ESBL-PE). The risk varies by destination and, for each traveller, may be substantially further increased by travellers' diarrhoea (TD) and antibiotic use. Despite the risk of TD in Africa, ESBL-PE acquisition rates in all studies are lower there than in Asia. Africa has become increasingly popular as a destination for international travellers, yet minimal data are available from the continent's subregions and countries., Methods: We analysed subregion- and country-specific data on carriage and risk factors for ESBL-PE colonization pooled from three prospective studies conducted between 2009 and 2013 among Finnish and Dutch travellers. The data were subjected to multivariable analysis of risk factors. In addition, we compared our data to two recent large investigations reporting data by subregion and country., Results: Our joint analysis comprised data on 396 travellers. The ESBL-PE colonization rate was highest in Northern Africa, followed by Middle and Eastern Africa, and lowest in Southern and Western Africa. Of individual countries with more than 15 visitors, the highest rates were seen for Egypt (12/17; 70.6%), Ghana (6/23; 26.1%), and Tanzania (14/81; 17.3%); the rates among travellers to Egypt were comparable to those reported in South and Southeast Asia. In a pooled multivariable analysis, travel destination, age, overnight hospitalisation abroad, TD, and use of fluoroquinolones were independently associated with increased ESBL-PE colonization rates., Conlusions: Even in areas with relatively low risk of colonization, antimicrobials clearly predispose to colonization with ESBL-PE. Travellers to Africa should be cautioned against unnecessary use of antibiotics.

A 21-year-old man had painful lesions on his scrotum for 4 days. Recently, bed bugs were discovered in the student accommodation of his girlfriend. We also observed vesicular lesions on the patient's buttock. PCR on swab specimens was negative for herpes simplex virus type 1 and 2, but positive for varicella zoster virus.

Background: Lyme borreliosis (LB) is a tick-borne infection caused by Borrelia burgdorferi sensu lato. The most frequent clinical manifestations are erythema migrans and Lyme neuroborreliosis. Currently, a large volume of diagnostic testing for LB is reported, whereas the incidence of clinically relevant disease manifestations is low. This indicates overuse of diagnostic testing for LB with implications for patient care and cost-effective health management., Aim: The recommendations provided in this review are intended to support both the clinical diagnosis and initiatives for a more rational use of laboratory testing in patients with clinically suspected LB., Sources: This is a narrative review combining various aspects of the clinical and laboratory diagnosis with an educational purpose. The literature search was based on existing systematic reviews, national and international guidelines and supplemented with specific citations., Implications: The main recommendations according to current European case definitions for LB are as follows. Typical erythema migrans should be diagnosed clinically and does not require laboratory testing. The diagnosis of Lyme neuroborreliosis requires laboratory investigation of the spinal fluid including intrathecal antibody production, and the remaining disease manifestations require testing for serum antibodies to B. burgdorferi. Testing individuals with non-specific subjective symptoms is not recommended, because of a low positive predictive value., (Copyright © 2017 European Society of Clinical Microbiology and Infectious Diseases. All rights reserved.)

Background: Control of gonorrhea in resource-limited countries, such as Indonesia, is mostly unsuccessful. Examining Neisseria gonorrhoeae (Ng) transmission networks using strain typing might help prioritizing public health interventions., Methods: In 2014, urogenital Ng strains were isolated from clients of sexually transmitted infection clinics in three Indonesian cities. Strains were typed using Multiple-Locus Variable Number Tandem Repeat (VNTR) Analysis (MLVA) and Ng Multi-Antigen Sequence Typing (NG-MAST) at the Public Health Service, Amsterdam, the Netherlands, and compared to Dutch strains collected from 2012 to 2015. Minimum spanning trees (MSTs) were constructed using MLVA profiles incorporating demographics and NG-MAST genogroups. A cluster was defined as ≥5 strains differing in ≤1 VNTR locus. The concordance between MLVA and NG-MAST was examined with the adjusted Wallace coefficients (AW)., Results: We collected a total of 78 Indonesian strains from Yogyakarta (n = 44), Jakarta (n = 25), and Denpasar (n = 9). Seven MLVA clusters and 16 non-clustered strains were identified. No cluster was specific for any geographic area, risk group or age group. Combined with 119 contemporary Dutch strains, 8 MLVA clusters were identified, being four clusters of Indonesian strains, two of Dutch strains, and two of both Indonesian and Dutch strains. Indonesian strains (79.5%) were more often clustered compared to Dutch strains (24.3%). The most prevalent NG-MAST genogroups among Indonesian strains was G1407 (51.3%) and among Dutch strains was G2992 (19.3%). In Indonesian strains, the AW [95% confidence interval] for MLVA to NG-MAST was 0.07 [0.00-0.27] and for NG-MAST to MLVA was 0.03 [0.00-0.12]., Conclusion: Indonesian Ng strains are more often clustered than Dutch strains, but show no relation with geographical area, risk group, or age group, suggesting a more clonal Ng epidemic in Indonesia. Some similar Ng strains circulate in both Indonesia and the Netherlands.

Introduction: Infection of Neisseria gonorrhoeae in the pharynx (pharyngeal Ng) is associated with gonococcal transmission and development of antimicrobial resistance. We assessed proportion of and determinants for persistence after treatment of pharyngeal Ng., Methods: At the STI clinic of Amsterdam, the Netherlands, females-at-risk and men who have sex with men are routinely screened for pharyngeal Ng using an RNA-based nucleic acid amplification test (NAAT; Aptima Combo 2). Patients with pharyngeal Ng were invited for a test-of-cure (TOC) 7 days after treatment with a 500 mg ceftriaxone intramuscularly. We retrospectively examined medical records of patients with pharyngeal Ng (January 2012-August 2015) who returned for a TOC 7-28 days after treatment. Persistence was defined as a positive NAAT at TOC., Results: Out of 2204 pharyngeal Ng cases recorded in the study period, 781 cases (median time between first treatment and TOC of 8 (IQR 7-12) days) were included in the analysis. Persistence after treatment was found in 36 (4.6%) and was less likely among patients who received ceftriaxone in combination with other antibiotics (vs monotherapy) (adjusted OR (aOR) 0.36, 95% CI 0.12 to 1.04) and with longer time from treatment to TOC (aOR 0.74, 95% CI 0.60 to 0.90, per extra day). In those with a TOC 15-28 days after treatment, Ng persisted in only 1.0% (1/105 cases)., Conclusion: A small proportion of pharyngeal Ng persists despite appropriate treatment. Combining ceftriaxone with other antibiotics appears to lead to faster clearance. A TOC for pharyngeal Ng 7 days after treatment may be too soon., Competing Interests: Competing interests: HJCDV and MFSVL served on a vaccine advisory board of GSK. Other authors have no conflict of interest to declare., (© Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.)