Achyuta Nori

Rectal chlamydia - a reservoir of undiagnosed infection in men who have sex with men.

Objective: To determine the prevalence of rectal chlamydia infection in a cohort of men who have sex with men (MSM) and the proportion of infection that would be missed without routine screening. Methods: MSM presenting to four HIV/GUM outpatient clinics at the Chelsea & Westminster Hospital NHS Foundation Trust between 1 November 2005 and 29 September 2006 were offered testing for rectal chlamydia infection in addition to their routine screen for sexually transmitted infections (STIs). Chlamydia trachomatis (CT) tests were performed using the Beckton-Dickinson Probe-Tec Strand Displacement Assay. Positive samples were re-tested at the Sexually Transmitted Bacteria Reference Laboratory, to confirm the result and identify lymphogranuloma venereum (LGV)-associated serovars. Results: A total of 3076 men were screened. We found an 8.2% prevalence of infection with CT (LGV and non-LGV serovars) in the rectum and 5.4% in the urethra. The HIV and rectal chlamydia co-infection rate was 38.1%. The majority of rectal infections (69.2%, (171/247)) were asymptomatic and would have been missed if routine screening had not been undertaken. Of the samples re-tested, 94.2% (227/242) rectal and 91.8% (79/86) urethral specimens were confirmed CT positive and 36 cases of LGV were identified. Conclusion: Our data show a high rate of rectal chlamydia infection, in the majority of cases it was asymptomatic. We recommend routine screening for rectal chlamydia in men at risk, as this may represent an important reservoir for the onward transmission of infection.

High Prevalence of Antibiotic-Resistant Mycoplasma genitalium in Nongonococcal Urethritis: The Need for Routine Testing and the Inadequacy of Current Treatment Options

Mycoplasma genitalium infections were as frequent a cause of nongonococcal urethritis as Chlamydia trachomatis, had high rates of macrolide-associated genotypic resistance, and were nonclonal, suggesting an established community infection. Detection of genotypic resistance to fluoroquinolones is cause for concern.

Performance evaluation of automated urine microscopy as a rapid, non-invasive approach for the diagnosis of non-gonococcal urethritis

Objectives Gram-stained urethral smear (GSUS), the standard point-of-care test for non-gonococcal urethritis (NGU) is operator dependent and poorly specific. The performance of rapid automated urine flow cytometry (AUFC) of first void urine (FVU) white cell counts (UWCC) for predicting Mycoplasma genitalium and Chlamydia trachomatis urethral infections was assessed and its application to asymptomatic infection was evaluated. Methods Receiver operating characteristic curve analysis, determining FVU-UWCC threshold for predicting M. genitalium or C. trachomatis infection was performed on 208 ‘training’ samples from symptomatic patients and subsequently validated using 228 additional FVUs obtained from prospective unselected patients. Results An optimal diagnostic threshold of >29 UWC/µL gave sensitivities and specificities for either infection of 81.5% (95% CI 65.1% to 91.6%) and 85.8% (79.5% to 90.4%), respectively, compared with 86.8% (71.1% to 95%) and 64.7% (56.9% to 71.7%), respectively, for GSUS, using the training set samples. FVU-UWCC demonstrated sensitivities and specificities of 69.2% (95% CI 48.1% to 84.9%) and 92% (87.2% to 95.2%), respectively, when using validation samples. In asymptomatic patients where GSUS was not used, AUFC would have enabled more infections to be detected compared with clinical considerations only (71.4% vs 28.6%; p=0.03). The correlation between UWCC and bacterial load was stronger for M. genitalium compared with C. trachomatis (τ=0.426, p≤0.001 vs τ=0.295, p=0.022, respectively). Conclusions AUFC offers improved specificity over microscopy for predicting C. trachomatis or M. genitalium infection. Universal AUFC may enable non-invasive diagnosis of asymptomatic NGU at the PoC. The degree of urethral inflammation exhibits a stronger association with pathogen load for M. genitalium compared with C. trachomatis.

Impact of deploying multiple point-of-care tests with a ‘sample first’ approach on a sexual health clinical care pathway. A service evaluation

Objectives To assess clinical service value of STI point-of-care test (POCT) use in a ‘sample first’ clinical pathway (patients providing samples on arrival at clinic, before clinician consultation). Specific outcomes were: patient acceptability; whether a rapid nucleic acid amplification test (NAAT) for Chlamydia trachomatis/Neisseria gonorrhoeae (CT/NG) could be used as a POCT in practice; feasibility of non-NAAT POCT implementation for Trichomonas vaginalis (TV) and bacterial vaginosis (BV); impact on patient diagnosis and treatment. Methods Service evaluation in a south London sexual health clinic. Symptomatic female and male patients and sexual contacts of CT/NG-positive individuals provided samples for diagnostic testing on clinic arrival, prior to clinical consultation. Tests included routine culture and microscopy; CT/NG (GeneXpert) NAAT; non-NAAT POCTs for TV and BV. Results All 70 (35 males, 35 females) patients approached participated. The ‘sample first’ pathway was acceptable, with >90% reporting they were happy to give samples on arrival and receive results in the same visit. Non-NAAT POCT results were available for all patients prior to leaving clinic; rapid CT/NG results were available for only 21.4% (15/70; 5 males, 10 females) of patients prior to leaving clinic. Known negative CT/NG results led to two females avoiding presumptive treatment, and one male receiving treatment directed at possible Mycoplasma genitalium infection causing non-gonococcal urethritis. Non-NAAT POCTs detected more positives than routine microscopy (TV 3 vs 2; BV 24 vs 7), resulting in more patients receiving treatment. Conclusions A ‘sample first’ clinical pathway to enable multiple POCT use was acceptable to patients and feasible in a busy sexual health clinic, but rapid CT/NG processing time was too long to enable POCT use. There is need for further development to improve test processing times to enable POC use of rapid NAATs.

Genomic epidemiology of syphilis reveals independent emergence of macrolide resistance across multiple circulating lineages

Syphilis is an ancient sexually transmitted infection caused by the bacterium Treponema pallidum subspecies pallidum and may lead to severe clinical complications. Recent years have seen striking increases in syphilis diagnoses in many high income countries, with the UK reporting a 148% increase in new diagnoses over 10 years. The reasons for this rise are complex and multifactorial, including changing cultural, behavioural, and technological factors that influence sexual networks and transmission dynamics. Previous genomic analyses have suggested that one lineage of syphilis, called SS14, may have expanded recently, with most syphilis caused by this lineage, and that this expansion indicates emergence of a single pandemic azithromycin-resistant cluster. In this study, we used high throughput sequencing of Treponema pallidum performed on DNA extracted directly from clinical swab samples and clinically derived samples with minimal passage in the rabbit to more than double the number of publicly available whole genome sequences. We used phylogenomic and population genomic analyses to show that both SS14-lineage and Nichols-lineage T. pallidum are present in contemporary patients and that SS14 is polyphyletic lineage. We further correlate the appearance of genotypic macrolide resistance with multiple SS14 sub-lineages, showing that both genotypically macrolide resistant and macrolide sensitive sub-lineages are spreading contemporaneously. These findings demonstrate that macrolide resistance has independently evolved multiple times in T. pallidum, that once evolved it becomes fixed in the genome and is transmissible, and tha these findings are not consistent with the hypothesis of SS14-lineage expansion purely due to macrolide resistance. Beyond relevance to our understanding of the current syphilis epidemic, these findings show how macrolide resistance evolves in Treponema subspecies. Furthermore, the evolution of macrolide resistance, despite not being first-line treatment, provides a warning on broader issues of antimicrobial resistance, and highlights the importance of stewardship and strategic planning to prevent the emergence of antimicrobial resistance.

Diagnostic accuracy of a prototype rapid chlamydia and gonorrhoea recombinase polymerase amplification assay: a multicentre cross-sectional preclinical evaluation

Objectives Rapid and accurate sexually transmitted infection diagnosis can reduce onward transmission and improve treatment efficacy. We evaluated the accuracy of a 15-minute run-time recombinase polymerase amplification–based prototype point-of-care test (TwistDx) for Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG). Methods Prospective, multicentre study of symptomatic and asymptomatic patients attending three English sexual health clinics. Research samples provided were additional self-collected vulvovaginal swab (SCVS) (female participants) and first-catch urine (FCU) aliquot (female and male participants). Samples were processed blind to the comparator (routine clinic CT/NG nucleic acid amplification test (NAAT)) results. Discrepancies were resolved using Cepheid CT/NG GeneXpert. Results Both recombinase polymerase amplification and routine clinic NAAT results were available for 392 male and 395 female participants. CT positivity was 8.9% (35/392) (male FCU), 7.3% (29/395) (female FCU) and 7.1% (28/395) (SCVS). Corresponding NG positivity was 3.1% (12/392), 0.8% (3/395) and 0.8% (3/395). Specificity and positive predictive values were 100% for all sample types and both organisms, except male CT FCU (99.7% specificity (95% confidence interval (CI) 98.4–100.0; 356/357), 97.1% positive predictive value (95% CI 84.7–99.9; 33/34)). For CT, sensitivity was ≥94.3% for FCU and SCVS. CT sensitivity for female FCU was higher (100%; 95% CI, 88.1–100; 29/29) than for SCVS (96.4%; 95% CI, 81.7–99.9; 27/28). NG sensitivity and negative predictive values were 100% in FCU (male and female). Conclusions This prototype test has excellent performance characteristics, comparable to currently used NAATs, and fulfils several World Health Organization ASSURED criteria. Its rapidity without loss of performance suggests that once further developed and commercialized, this test could positively affect clinical practice and public health.

P015 Receiving 1g azithromycin as part of mass drug administration (mda) for the control of trachoma is associated with reduced genital mycoplasma genitaliumprevalence

Introduction Mass Drug Administration (MDA) with 1g oral azithromycin for ocular Chlamydia trachomatis (CT) infection, a key component of trachoma control, can concomitantly reduce genital CT prevalence. However, this dose is known to be sub-optimal for the treatment of genital Mycoplasma genitalium (MG) infection. Here we investigate factors associated with MG infection in pre- and post-MDA sample sets. Methods Pre-MDA (T1) and 6 months post-MDA (T2) CT-negative self-collected vulvo-vaginal swabs from women attending three outpatient antenatal clinics (Honiara, Solomon Islands), were tested for MG infection using nucleic acid amplification. Logistic regression was used to determine factors associated with infection. Variables tested included: patient age, clinic attended, ethnicity, time spent in education, living in an urban or rural environment, marital status, living with spouse, presence of symptoms associated with a sexually transmitted infection (STI), having an STI in the last 12 months, current CT, Gonorrhoea or Trichomonas vaginalis infection, and at T2 only receipt of MDA dose. Results MG positivity was found in 11.9% (95%CI: 8.3–16.6; 28/236) of women at T1 and in 10.9% (95%CI: 7.7–15.4; 28/256) at T2 (p=0.7467). The only factor associated with having an MG infection was history of not having received MDA with azithromycin at T2 (odds ratio 0.19, 95%CI 0.07–0.53, p=0.001). Discussion Not having MG infection was associated with receiving 1g azithromycin as part of MDA for trachoma control six months previously. However there was no overall drop in population prevalence, indicating individual but not population benefits of MDA with regard to MG infection control.

P1.38 Bacterial populations detected within first void urine samples of symptomatic male patients with urethritis

Introduction Applying bacterial 16S rRNA profiling we investigated whether species in first void urine (FVU) differed between men presenting with urethritis symptoms with and without urethral inflammation. Methods 443 patients prospectively attending a London sexual health clinic were classified, based on clinical presentation and Gram stain as: symptomatic urethritis (URE); symptomatic non-urethritis (SYM); and asymptomatic (ASP). Residual FVU’s were tested for [K1] Chlamydia trachomatis (CT), Neisseria gonorrhoeae (NG), Trichomonas vaginalis (TV) and Mycoplasma genitalium (MG) and subjected to bacterial 16S rRNA profiling (variable region v1-2) Differences in bacterial oligotypes across samples were described using partial least square discriminant analysis tested using permutational multivariate analysis of variance (PERMANOVA). The hypothesis tested was: URE have distinct urinary bacterial 16S rRNA profiles compared to SYM and ASP. Results 286/443 samples met quality control criteria. Among URE [n=79], SYM [n=83], and ASP [n=124], CT, MG, NG and TV prevalence’s were: 12.7% (95 CI 6.6–22.5), 13.9% (95CI 7.5–24), 8.9% (95CI 3.9–7.5), 1.3% (95CI 0.1–7.8); 2.4% (95CI 0.4–9.2), 2.4% (95CI 0.4–9.2), 1.2% (95CI 0.1–7.5); 2.4% (95CI 0.6–7.4), 5.7% (95CI 2.5–11.8), NG not detected, 0.8% (95CI 0–5.1) respectively. Lactobacillus iners was the most abundant oligotype observed in all groups. Streptococcus agalactiae and S. anginosus were highly dominant in ASP along with S. mitis, which was relatively increased in both ASP and SYM. A decreased relative abundance of these oligotypes was observed in URE cases. A significant difference in oligotype populations was observed between URE and SYM (PERMANOVA test p=0.0072, F2.182), but not between SYM and ASP (PERMANOVA test [JW2] p=0.0.1797, F1.601). Conclusion SYM and ASP oligotype populations were dominated by Lactobacillus and Streptococcus species. Microbiota observed in FVU samples of symptomatic patients with microscopy confirmed urethritis (URE) were distinct to those with symptoms but no urethritis on microscopy (SYM).

P1.012 Urinary Cytokine Profiles in Non-Specific, Mycoplasma Genitalium and Chlamydia Trachomatis Urethritis

Background Previously, we demonstrated that urinary white cell count increases in proportion to pathogen load in cases of urethritis caused by Mycoplasma genitalium but not Chlamydia trachomatis. We further investigated urethritis pathogenesis caused by these organisms by comparing concentrations of 23 cytokines present within first void urine (FVU) specimens of male urethritis cases. Methods FVUs from 52 symptomatic male patients (all underwent Gram stain urethral smear) were collected and patients stratified into those with non-specific urethritis (n = 12), M. genitalium urethritis (n = 13), C. trachomatis urethritis (n = 14) and non-urethritis controls (n = 13). Cytokines measurements from FVUs specimens were obtained using a Human 30-Plex Luminex assay. Concentrations were obtained for 23 of the 30 cytokines analysed and compared between the four groups using multivariate ANOVA. Results Overall, there was a significant difference in urinary cytokine profile between groups (p = 0.042). For individual cytokines, clinical group was associated with differences in concentrations of IL-1β (p = 0.007), GCSF (p = 0.042), CCL11 (p = 0.012), MIP-1α (p = 0.029), TNF-α (p = 0.026), IL-7 (p = 0.029), EGF (p = 0.030), VEGF (p = 0.049) and IFNa (p = 0.008). When compared with uninfected non-urethritis controls, cytokine concentrations in: M. genitalium samples, were increased for IL-1β (p = 0.017), GCSF (p = 0.010) but decreased for EGF (p = 0.017); C. trachomatissamples, were decreased for EGF (p = 0.049); and in non-specific urethritis samples, were increased for CCL5 (p = 0.049), IL-1β (p = 0.05), IL-1RA (p = 0.033) and decreased for EGF (p = 0.032). No significant differences were demonstrated in cytokine concentrations between C. trachomatis and M. genitalium groups. Conclusion The increased levels of pro-inflammatory cytokines present in the urethritis groups when compared to non-urethritis controls reflect the acute inflammatory state. The data suggests that M. genitaliumgenital infection may be associated with a discrete mucosal immunological profile potentially explaining the link between cellular inflammatory response and bacterial load, previously observed