Dinner date: Neisseria gonorrhoeae central carbon metabolism and pathogenesis.

Neisseria gonorrhoeae, the causative agent of the sexually transmitted infection gonorrhea, is a human-adapted pathogen that does not productively infect other organisms. The ongoing relationship between N. gonorrhoeae and the human host is facilitated by the exchange of nutrient resources that allow for N. gonorrhoeae growth in the human genital tract. What N. gonorrhoeae 'eats' and the pathways used to consume these nutrients have been a topic of investigation over the last 50 years. More recent investigations are uncovering the impact of N. gonorrhoeae metabolism on infection and inflammatory responses, the environmental influences driving N. gonorrhoeae metabolism, and the metabolic adaptations enabling antimicrobial resistance. This mini-review is an introduction to the field of N. gonorrhoeae central carbon metabolism in the context of pathogenesis. It summarizes the foundational work used to characterize N. gonorrhoeae central metabolic pathways and the effects of these pathways on disease outcomes, and highlights some of the most recent advances and themes under current investigation. This review ends with a brief description of the current outlook and technologies under development to increase understanding of how the pathogenic potential of N. gonorrhoeae is enabled by metabolic adaptation.

Gonococcal invasion into epithelial cells depends on both cell polarity and ezrin.

Neisseria gonorrhoeae (GC) establishes infection in women from the cervix, lined with heterogeneous epithelial cells from non-polarized stratified at the ectocervix to polarized columnar at the endocervix. We have previously shown that GC differentially colonize and transmigrate across the ecto and endocervical epithelia. However, whether and how GC invade into heterogeneous cervical epithelial cells is unknown. This study examined GC entry of epithelial cells with various properties, using human cervical tissue explant and non-polarized/polarized epithelial cell line models. While adhering to non-polarized and polarized epithelial cells at similar levels, GC invaded into non-polarized more efficiently than polarized epithelial cells. The enhanced GC invasion in non-polarized epithelial cells was associated with increased ezrin phosphorylation, F-actin and ezrin recruitment to GC adherent sites, and the elongation of GC-associated microvilli. Inhibition of ezrin phosphorylation inhibited F-actin and ezrin recruitment and microvilli elongation, leading to a reduction in GC invasion. The reduced GC invasion in polarized epithelial cells was associated with non-muscle myosin II-mediated F-actin disassembly and microvilli denudation at GC adherence sites. Surprisingly, intraepithelial GC were only detected inside epithelial cells shedding from the cervix by immunofluorescence microscopy, but not significantly in the ectocervical and the endocervical regions. We observed similar ezrin and F-actin recruitment in exfoliated cervical epithelial cells but not in those that remained in the ectocervical epithelium, as the luminal layer of ectocervical epithelial cells expressed ten-fold lower levels of ezrin than those beneath. However, GC inoculation induced F-actin reduction and myosin recruitment in the endocervix, similar to what was seen in polarized epithelial cells. Collectively, our results suggest that while GC invade non-polarized epithelial cells through ezrin-driven microvilli elongation, the apical polarization of ezrin and F-actin inhibits GC entry into polarized epithelial cells.

Transcriptional and Translational Responsiveness of the Neisseria gonorrhoeae Type IV Secretion System to Conditions of Host Infections.

The type IV secretion system of Neisseria gonorrhoeae translocates single-stranded DNA into the extracellular space, facilitating horizontal gene transfer and initiating biofilm formation. Expression of this system has been observed to be low under laboratory conditions, and multiple levels of regulation have been identified. We used a translational fusion of lacZ to traD, the gene for the type IV secretion system coupling protein, to screen for increased type IV secretion system expression. We identified several physiologically relevant conditions, including surface adherence, decreased manganese or iron, and increased zinc or copper, which increase gonococcal type IV secretion system protein levels through transcriptional and/or translational mechanisms. These metal treatments are reminiscent of the conditions in the macrophage phagosome. The ferric uptake regulator, Fur, was found to repress traD transcript levels but to also have a second role, acting to allow TraD protein levels to increase only in the absence of iron. To better understand type IV secretion system regulation during infection, we examined transcriptomic data from active urethral infection samples from five men. The data demonstrated differential expression of 20 of 21 type IV secretion system genes during infection, indicating upregulation of genes necessary for DNA secretion during host infection.

Lactiplantibacillus plantarum strains isolated from spontaneously fermented cocoa exhibit potential probiotic properties against Gardnerella vaginalis and Neisseria gonorrhoeae.

BACKGROUND: Probiotics are important tools in therapies against vaginal infections and can assist traditional antibiotic therapies in restoring healthy microbiota. Recent research has shown that microorganisms belonging to the genus Lactobacillus have probiotic potential. Thus, this study evaluated the potential in vitro probiotic properties of three strains of Lactiplantibacillus plantarum, isolated during the fermentation of high-quality cocoa, against Gardnerella vaginalis and Neisseria gonorrhoeae. Strains were evaluated for their physiological, safety, and antimicrobial characteristics. RESULTS: The hydrophobicity of L. plantarum strains varied from 26.67 to 91.67%, and their autoaggregation varied from 18.10 to 30.64%. The co-aggregation of L. plantarum strains with G. vaginalis ranged from 14.73 to 16.31%, and from 29.14 to 45.76% with N. gonorrhoeae. All L. plantarum strains could moderately or strongly produce biofilms. L. plantarum strains did not show haemolytic activity and were generally sensitive to the tested antimicrobials. All lactobacillus strains were tolerant to heat and pH resistance tests. All three strains of L. plantarum showed antimicrobial activity against the tested pathogens. The coincubation of L. plantarum strains with pathogens showed that the culture pH remained below 4.5 after 24 h. All cell-free culture supernatants (CFCS) demonstrated activity against the two pathogens tested, and all L. plantarum strains produced hydrogen peroxide. CFCS characterisation in conjunction with gas chromatography revealed that organic acids, especially lactic acid, were responsible for the antimicrobial activity against the pathogens evaluated. CONCLUSION: The three strains of L. plantarum presented significant probiotic characteristics against the two pathogens of clinical importance. In vitro screening identified strong probiotic candidates for in vivo studies for the treatment of vaginal infections.

Human endometrial MAIT cells are transiently tissue resident and respond to Neisseria gonorrhoeae.

Mucosa-associated invariant T (MAIT) cells are non-classical T cells important in the mucosal defense against microbes. Despite an increasing interest in the immunobiology of the endometrial mucosa, little is known regarding human MAIT cells in this compartment. The potential role of MAIT cells as a tissue-resident local defense against microbes in the endometrium is largely unexplored. Here, we performed a high-dimensional flow cytometry characterization of MAIT cells in endometrium from pre- and postmenopausal women, and in decidua from first-trimester pregnancies. Furthermore, we assessed MAIT cell function by stimulation with Neisseria gonorrhoeae (N. gonorrhoeae). Endometrial MAIT (eMAIT) cells represented a stable endometrial immune cell population as limited dynamic changes were observed during the menstrual cycle, post menopause, or in response to pregnancy. Furthermore, eMAIT cells exhibited an activated tissue-resident phenotype. Despite expressing CD69 and CD103, eMAIT cells were replenished over time by circulating MAIT cells, as assessed using human uterus transplantation as a model. Finally, functional experiments revealed the capability of MAIT cells to respond to the sexually transmitted and tissue-relevant pathogen, N. gonorrhoeae. In conclusion, our study provides novel insight into human MAIT cell dynamics and anti-microbial properties in the human uterus.

Theoretical modelling of competitive microbial range expansion with heterogeneous mechanical interactions.

Microbial range expansion experiments provide insight into the complex link between dynamic structure, pattern formation and evolutionary dynamics of growing populations. In this work, we develop a theoretical model in order to investigate the interplay of growth statistics and mechanical interactions which are implemented as division driven pushing and swapping of cells. For the case of the competitive growth of a strongly and a weakly interacting strain we investigate the influence of different mean division times, as well as different mechanical interactions on the development of the colony. Our results show that the susceptibility to cell division induced pushing has a much stronger influence on the structure of the colony than cell sorting towards the colony's perimeter. Motivated by microbial range expansion experiments of Neisseria gonorrhoeae bacteria, we also consider the influence of mutating cells on the structure of the colony. We show that the outgrowth of the three different strains is strongly influenced by the relative strengths of their mechanical interaction. The experimentally observed patterns are reproduced for mechanical interactions of the mutants, which range between those of the strongly and weakly interacting strain.

Antibacterial Activity of Chitosan Nanoparticles Against Pathogenic N. gonorrhoea.

PURPOSE: The emergence of Neisseria gonorrhoeae strains that are resistant to the most commonly used antibiotics represents a great concern for global public health. This challenges the effectiveness of clinical treatment regimens and demands the development of alternative antigonococcal agent. In this regard, chitosan nanoparticles (CNPs) are known to have antimicrobial activity against a wide range of pathogens. Thus, they have become a potential candidate for combatting this era of multi-drug resistance. This study aims to formulate CNPs, characterize their physicochemical properties, and examine their antimicrobial activity against gonococcus. MATERIALS AND METHODS: The ionic gelation method was used to prepare CNPs of different concentrations. Characterization for their particle size (PZ), polydispersity index (PDI), and zeta potential (ZP) was performed. The anti-microbial activity of CNPs was investigated against 13 WHO N. gonorrhoeae reference strains, using the broth dilution method. Cytotoxicity of CNPs and their effect on bacterial adhesion to HeLa cells were investigated. RESULTS: The average PZ and ZP of the prepared NPs were increased when the concentration of chitosan was increased from 1 to 5 mg/mL and found to be in the range of 193 nm ± 1.9 to 530 nm ± 13.3, and 14 mV ± 0.5 to 20 mV ± 1, respectively. Transmission electron microscopes (TEM) images revealed spherical NPs, and the NPs had a low PDI value of ≤0.27. The formed CNPs produced antibacterial activity against all tested strains, including those resistant to multiple antibiotics, with a minimum inhibitory concentration (MIC90) of 0.16 to 0.31 mg/mL and a minimum bactericidal concentration (MBC) of 0.31 to 0.61 mg/mL. Of note, at all MIC90 and MBC, the CNPs had no significant cytotoxic effect on HeLa cells and reduced bacterial adhesion to these cells at MBC doses. CONCLUSION: The present work findings suggest the potential of the CNPs for the treatment of gonorrhoea.

Oral sex practices among men who have sex with men and transgender women at risk for and living with HIV in Nigeria.

BACKGROUND: Men who have sex with men (MSM) and transgender women (TGW) are at risk for sexually transmitted infections (STIs), including those of the oropharynx. We estimated the prevalence and factors associated with oral sex practices and characterized oropharyngeal STIs among a cohort of MSM and TGW in Nigeria. METHODS: From 2013 to 2018, TRUST/RV368 recruited MSM and TGW into HIV/STI diagnosis and treatment at community-based clinics in Nigeria. Participants who completed HIV testing and oral sex questions at enrollment were selected. Cross-sectional analyses with bivariate and multivariable logistic regression models estimated odds ratios (ORs) and 95% confidence intervals (CIs). Oropharyngeal swab testing for Neisseria gonorrhoeae (NG) and Chlamydia trachomatis (CT) began in 2014 and for those with diagnostic results at enrollment, the unadjusted association of oral sex practices with oropharyngeal STIs was conducted. RESULTS: A total of 1342 participants had a median age of 25 years (interquartile range: 22-29), 58% were living with HIV, and 69% reported oral sex practices. Factors associated with increased odds of engaging in oral sex included living with HIV (adjusted [a]OR: 1.4, 95% CI: 1.1-1.8), self-identifying as a woman (aOR:1.8, 95% CI: 1.1-2.8), mobile phone ownership (aOR:2.3, 95% CI: 1.3-3.9), receptive anal sex (aOR:1.7, 95% CI:1.3-2.3) and multiple male sexual partners (2 to 4 vs. ≤1, aOR:1.5, 95% CI: 1.0-2.2; 5+ vs ≤1, aOR:2.9, 95% CI:1.9-4.3). Oropharyngeal STI prevalence was 7% (52/752) and higher among those who engaged in oral sex compared to those who did not (unadjusted OR: 2.5, 95% CI:1.2-5.3). CONCLUSIONS: Oral sex was common and associated with an increased odds of oropharyngeal STIs among MSM and TGW from Nigeria. In the absence of screening and treatment guidelines, condoms continue to be the mainstay for oral STI prevention. A pre-exposure prophylaxis for bacterial STIs would complement current prevention strategies to curb transmission.

Spontaneous clearance of genital and extragenital Neisseria gonorrhoeae: data from GToG.

OBJECTIVES: Neisseria gonorrhoeae (NG) infection can resolve without antibiotic treatment, however the literature describing the frequency of clearance at individual sites, how rapidly it occurs and potential predictive factors is limited. In this analysis of a subpopulation identified from a large multicentre UK cohort, we describe the overall rate of spontaneous clearance of infection and explore factors associated with this. METHODS: Data from the Gentamicin compared with Ceftriaxone for the Treatment of Gonorrhoea randomised controlled trial consisting of 720 patients with NG were analysed. A subgroup of individuals had both a pretrial test sample and a trial enrolment sample taken. Those who had cleared NG between initial presentation and subsequent entry into the trial without antibiotic treatment were deemed to have spontaneously cleared. Sociodemographic characteristics, sexual history and sites of infection for those who spontaneously cleared infection were compared with that of those who did not. We also estimated the time interval to clearance. RESULTS: Overall, the proportion who had spontaneous clearance was 20.5% (83/405). Clearance of infection occurred over a median of 10 days (IQR 7-15 days). The cohort who spontaneously cleared were similar to those who did not in terms of age, gender, sexual orientation, HIV status and previous NG infection. Chlamydia coinfection was more frequent in the 'no spontaneous clearance group' (11.1% (9/83) cf 22.0% (69/322)) (p=0.029). Dysuria was reported more often in the 'no spontaneous clearance group' (4.8% (4/83) cf 13.0% (42/322)) (p=0.035). CONCLUSION: We present data from a large cohort of NG-infected individuals, of whom a significant proportion had spontaneous clearance of infection. This is consistent with previous smaller studies. If this is indicative of cure, point-of-care testing prior to treatment has the potential to reduce unnecessary exposure to antimicrobials. Further work to assess the importance of bacterial load, genotype and host immune response on spontaneous clearance of infection is required. TRIAL REGISTRATION NUMBER: ISRCTN51783227.

Cross-sectional study of asymptomatic Neisseria gonorrhoeae and Chlamydia trachomatis infections in sexually transmitted disease related clinics in Shenzhen, China.

The aims of this study were to investigate the prevalence and proportion of laboratory-confirmed urethral Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) infections that were asymptomatic among individuals presenting to clinics in Shenzhen and the risk factors related to asymptomatic CT infection. In a cross-sectional study, eligible individuals were invited to participate in the questionnaire, and urine specimens were collected to identify CT and NG infections using a nucleic acid amplification test (NAAT). Considering the differences in the presentation of symptoms between men and women, this study was stratified by gender. Corresponding outcomes were analyzed by Chi-square test and multivariate logistic regression. A total of 2,871 participants were asymptomatic and included in our analyses: 1120 (39.0%) men and 1751 (61.0%) women. The prevalence of asymptomatic NG and CT infections was 0.9% and 6.2% in men, and 0.4% and 7.9% in women, respectively. The proportion of asymptomatic urethral CT among men with urethral CT was 28.3%; for women, it was 34.2%. For asymptomatic men with CT, 3 independent risk factors were identified: (1) men under the age of 30 (aOR, 1.83; 95% CI, 1.11-3.03); (2) being employed in the commercial service work (2.82; 1.36-5.84); and (3) being recruited through the urological department (2.12; 1.19-3.79). For asymptomatic women with urethral CT, age less than 30 years was a risk factor. In conclusion, a substantial prevalence of asymptomatic CT infections was found among men and women presenting to clinics in Shenzhen. The significant correlation between asymptomatic CT infection and these risk factors could help identify high-risk populations and guide screening.

Lactobacillus crispatus and its enolase and glutamine synthetase influence interactions between Neisseria gonorrhoeae and human epithelial cells.

Neisseria gonorrhoeae, an obligatory human pathogen causes the sexually transmitted disease gonorrhea, which remains a global health problem. N. gonorrhoeae primarily infects the mucosa of the genitourinary tract, which in women, is colonized by natural microbiota, dominated by Lactobacillus spp., that protect human cells against pathogens. In this study, we demonstrated that precolonization of human epithelial cells with Lactobacillus crispatus, one of the most prevalent bacteria in the female urogenital tract, or preincubation with the L. crispatus enolase or glutamine synthetase impairs the adhesion and invasiveness of N. gonorrhoeae toward epithelial cells, two crucial steps in gonococcal pathogenesis. Furthermore, decreased expression of genes encoding the proinflam-matory cytokines, TNFα and CCL20, which are secreted as a consequence of N. gonorrhoeae infection, was observed in N. gonorrhoeae-infected epithelial cells that had been preco-lonized with L. crispatus or preincubated with enolase and glutamine synthetase. Thus, our results indicate that the protection of human cells against N. gonorrhoeae infection is a complex process and that L. crispatus and its proteins enolase and glutamine synthetase can have a potential role in protecting epithelial cells against gonococcal infection. Therefore, these results are important since disturbances of the micro-biota or of its proteins can result in dysbiosis, which is associated with increased susceptibility of epithelium to pathogens.

The Clinico-Epidemiological Profile of Patients with Gonorrhoea and Challenges in the Management of Neisseria gonorrhoeae Infection in an STI clinic, Ternopil, Ukraine (2013-2018).

Gonorrhea is the second most common sexually transmitted infection spreading worldwide and a serious public health problem. However, further data are required to improve the management of gonorrhea. Our aim was to review the features of gonococcal infection and characterize the challenges of its management. A retrospective descriptive study of the medical records of 136 adult patients with gonorrhea that visited Ternopil Regional Sexually Transmitted Infections Clinic (Ukraine) in 2013-2018 was performed. The male-to-female ratio was 6.6:1. Homosexually-acquired gonorrhoea was 3.7%. Also, most patients acquired gonorrhea in Ukraine (98.4%). The mean infectious period lasted 2-16 days, including the incubation period of 1-9 days and the period from the onset of symptoms to the first visit of the clinic of 1-7 days. The probability of N. gonorrhoeae transmission within the frame of the epidemiologic sexual chain was 1:2.4. Concurrent T. vaginalis (39.7%) and C. trachomatis (2.2%) were detected. HIV and syphilis screening rates were 1.6% and 0.7%, respectively. The examining rate of sexual partners was 11%, testing extragenital specimens - 0.7%, screening coverage for HIV - 46.3%, compliance with follow-up visits - 41.9%. Part of patients (16.2%) received monotherapy with clarithromycin, doxycycline, benzylpenicillin, azithromycin, or ofloxacin. The management of N. gonorrhoeae infections was compromised by a low rate of examining sexual partners, females and testing extragenital specimens, screening for HIV, compliance to follow-up visits, access to nucleic acid amplification tests, and receiving questionable or even obsolete antimicrobial treatment. Therefore, more accurate and comprehensive management of gonorrhea is urgently needed in Ukraine.

Ethnically diverse urban transmission networks of Neisseria gonorrhoeae without evidence of HIV serosorting.

OBJECTIVE: We aimed to characterise gonorrhoea transmission patterns in a diverse urban population by linking genomic, epidemiological and antimicrobial susceptibility data. METHODS: Neisseria gonorrhoeae isolates from patients attending sexual health clinics at Barts Health NHS Trust, London, UK, during an 11-month period underwent whole-genome sequencing and antimicrobial susceptibility testing. We combined laboratory and patient data to investigate the transmission network structure. RESULTS: One hundred and fifty-eight isolates from 158 patients were available with associated descriptive data. One hundred and twenty-nine (82%) patients identified as male and 25 (16%) as female; four (3%) records lacked gender information. Self-described ethnicities were: 51 (32%) English/Welsh/Scottish; 33 (21%) white, other; 23 (15%) black British/black African/black, other; 12 (8%) Caribbean; 9 (6%) South Asian; 6 (4%) mixed ethnicity; and 10 (6%) other; data were missing for 14 (9%). Self-reported sexual orientations were 82 (52%) men who have sex with men (MSM); 49 (31%) heterosexual; 2 (1%) bisexual; data were missing for 25 individuals. Twenty-two (14%) patients were HIV positive. Whole-genome sequence data were generated for 151 isolates, which linked 75 (50%) patients to at least one other case. Using sequencing data, we found no evidence of transmission networks related to specific ethnic groups (p=0.64) or of HIV serosorting (p=0.35). Of 82 MSM/bisexual patients with sequencing data, 45 (55%) belonged to clusters of ≥2 cases, compared with 16/44 (36%) heterosexuals with sequencing data (p=0.06). CONCLUSION: We demonstrate links between 50% of patients in transmission networks using a relatively small sample in a large cosmopolitan city. We found no evidence of HIV serosorting. Our results do not support assortative selectivity as an explanation for differences in gonorrhoea incidence between ethnic groups.

Characteristics and Impact of Disseminated Gonococcal Infection in the "Top End" of Australia.

The "Top End" of Australia is presently experiencing a gonorrhea epidemic. Gonococcal infection is usually limited to mucosal tissues but can lead to disseminated gonococcal infection (DGI), joint destruction, and severe sepsis. This study aimed to explore the epidemiology, presentation, management, and health-care impact of DGI in the Top End of the Northern Territory. Health records of patients diagnosed with proven, probable, or possible DGI between January 2010 and September 2018 were analyzed retrospectively. One hundred six cases of DGI were identified. Ninety-four patients (88.7%) were Indigenous Australian. The incidence of proven and probable DGI in the Indigenous population was 27.1 per 100,000 person-years, compared with 7.1 in the Top End population overall. Of 7,540 laboratory-proven gonococcal notifications, 1.3% (n = 97) were complicated by DGI. The highest incidence was in the 15-19-year age-group. Thirteen cases (12.3%) occurred in patients younger than 15 years. High rates of comorbid alcohol misuse, diabetes, systemic lupus erythematosus, rheumatic fever, and complement deficiency were observed. The "classic triad" of tenosynovitis, dermatitis, and polyarthralgia was rare. Ninety-four patients (88.7%) presented with purulent arthritis. Disseminated gonococcal infection was estimated to cause at least 10.0% of nonpenetrating septic arthritis in the Top End and 1,234 days of hospitalization during the study period. DGI is an important cause of morbidity in the Top End, particularly in the young, remote Indigenous Australian population. Clinical presentation varies from classical teaching. Urgent action in the health and community sector is required, particularly for at-risk populations, to prevent further debilitating and costly complications of gonococcal infection.

Antimicrobial Resistance in Neisseria gonorrhoeae and Treatment of Gonorrhea.

Gonorrhea and antimicrobial resistance (AMR) in Neisseria gonorrhoeae are major public health concerns globally. Dual antimicrobial therapy (mainly ceftriaxone 250-500 mg × 1 plus azithromycin 1-2 g × 1) is currently recommended in many countries. These dual therapies have high cure rates, have likely been involved in decreasing the level of cephalosporin resistance internationally, and inhibit the spread of AMR gonococcal strains. However, ceftriaxone-resistant strains are currently spreading internationally, predominately associated with travel to Asia. Furthermore, the first global treatment failure with recommended dual therapy was reported in 2016 and the first isolates with combined ceftriaxone resistance and high-level azithromycin resistance were reported in 2018 in the UK and Australia. New antimicrobials for treatment of gonorrhea are essential and, of the few antimicrobials in clinical development, zoliflodacin particularly appears promising. Holistic actions are imperative. These include an enhanced advocacy; prevention, early diagnosis, contact tracing, treatment, test-of-cure, and additional measures for effective management of anogenital and pharyngeal gonorrhea; antimicrobial stewardship; surveillance of infection, AMR and treatment failures; and intensified research, for example, regarding rapid molecular point-of-care detection of gonococci and AMR, novel AMR determinants, new antimicrobials, and an effective gonococcal vaccine, which is the only sustainable solution for management and control of gonorrhea.

Gonorrhoea and chlamydia in persons with HIV: number needed to screen.

OBJECTIVES: Current guidelines recommend screening sexually active persons with HIV (PWH) for Neisseria gonorrhoeae (NG) and Chlamydia trachomatis (CT) at least annually. Yet, screening rates in many HIV clinics remain low. In this study, we estimated the number needed to screen (NNS) to detect a NG and/or CT infection at each anatomic site among different subpopulations of PWH. NNS provides a concrete, practical measure to aid in assessing the practical impact of screening. METHODS : We included adults in care at three HIV Research Network sites in 2011-2014. Restricting to first tests within each year, annual NNS was defined as number of persons tested divided by number positive. We computed urogenital and extragenital NNS by age and risk group (women, men who have sex with women (MSW) and men who have sex with men (MSM)). RESULTS : A total of 16 864 NG/CT tests were included. Among patients aged ≤25 years, urogenital NNS was similar among women (15 (95% CI 6 to 71)), MSW (21 (95% CI 6 to 167)) and MSM (20 (95% CI 12 to 36)). Over 25, urogenital NNS increased to a greater extent for women (363 (95% CI 167 to 1000)) and MSW (160 (95% CI 100 to 333)) than MSM (46 (95% CI 38 to 56)). The increase for women versus MSM >25 remained significant (p<0.01) in multivariable analysis. Among MSM, rectal NNS was 5 (95% CI 3 to 7) and 10 (95% CI 9 to 12) for ≤25 and for >25 years and pharyngeal NNS values were 8 (95% CI 5 to 13) and 20 (95% CI 16 to 24). CONCLUSIONS: These findings suggest the importance of regular, at least annual NG/CT screening, particularly extragenital, of HIV positive MSM of all ages. They provide some support for age-based cutoffs for women and MSW (eg, universal screening for those aged ≤25 and targeted screening for those aged >25 years).