JOURNAL TRANSCRIPT
Original article
European Journal of Microbiology and Immunology 7 (2017) 3, pp. 176–186 DOI: 10.1556/1886.2017.00018
HIGH-THROUGHPUT TESTING OF UROGENITAL AND EXTRAGENITAL SPECIMENS FOR DETECTION OF CHLAMYDIA TRACHOMATIS AND NEISSERIA GONORRHOEAE WITH cobas® CT/NG Elizabeth M. Marlowe1, David Hardy2, Mark Krevolin2, Peter Gohl3, Alexander Bertram4, Rodney Arcenas1, Britta Seiverth5, Tanja Schneider5, Oliver Liesenfeld1,* 1
Medical and Scientific Affairs, Roche Molecular Diagnostics, Pleasanton, CA, USA Development, Roche Molecular Diagnostics, Inc., Pleasanton, CA, USA 3 Bioscientia, Ingelheim, Germany 4 Amedes MVZ Wagnerstibbe für Laboratoriumsmedizin, Hämostaseologie, Humangenetik und Mikrobiologie, Hannover, Germany 5 Development, Roche Molecular Diagnostics, Rotkreuz, Switzerland 2
Received: July 17, 2017; Accepted: August 14, 2017 We compared the analytical and clinical performance of cobas® CT/NG for use on the cobas® 6800/8800 Systems with the cobas® 4800 CT/NG Test from urogenital and extragenital specimens in over 12,000 specimens from both male and female subjects in Germany and the United States. The analytical sensitivity was ≤40 EB/ml for Chlamydia trachomatis (CT) and ≤1 CFU/ml for Neisseria gonorrhoeae (NG). Using clinical specimens, the overall percent agreement with the cobas® 4800 CT/NG Test was >98.5%. Across urogenital specimens, there were 93 discrepant specimens; 76 (93.8%) of 81 CT discrepant specimens were 6800+/4800− and 10 (83.3%) of 12 NG discrepant specimens were 6800+/4800−. Sequencing verified CT results for 45 (61.6%) of 73 samples positive by 6800 and 1 (20%) of 5 positive by 4800. Similarly, 7 (70.0%) of 10 NG samples positive by 6800 and 1 of 2 positive by 4800 were confirmed by sequencing. Among discrepant extragenital specimens (all 6800+/4800−), 7 (50%) of 14 oropharyngeal and 23 (76.7%) of 30 anorectal CT discordant samples were confirmed as CT positive by sequencing; all 8 anorectal and 20 (90.9%) of 22 oropharyngeal NG discordant results were also confirmed as NG positive. In conclusion, cobas® CT/NG for use on the cobas® 6800/8800 Systems provides high-throughput automated solutions for sexually transmitted infection (STI) screening programs. Keywords: cobas® CT/NG, Chlamydia trachomatis, Neisseria gonorrhoeae, molecular diagnostics, PCR, extragenital infection, genital infection
Introduction Infections with Chlamydia trachomatis (CT) and Neisseria gonorrhoeae (NG) are the leading bacterial causes of sexually transmitted infections (STIs) worldwide [1, 2]. Prevalence is highest in persons aged 15–24 years [3]. Left untreated, these infections can cause significant sequelae to sexual health. Infections with these organisms have been strongly associated with development of pelvic inflammatory disease, adverse pregnancy outcomes, and
an increased risk of human immunodeficiency virus (HIV) acquisition [3–7]. Many infections remain asymptomatic, with high numbers of infected patients who may not seek care and thus remaining a reservoir for infection. Many countries have screening or testing recommendations for CT and/or NG, particularly aimed at young women among whom the disease burden and the consequences of untreated infection are highest [8]. While control strategies for CT and NG focus on urogenital testing during the past decade, various studies from
* Corresponding author: Oliver Liesenfeld; Medical and Scientific Affairs, Roche Molecular Diagnostics, Pleasanton, CA, USA; Phone: +19257308006; Fax: +19257308985; E-mail:
[email protected] This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial 4.0 International License (https://creativecommons.org/licenses/by-nc/4.0/), which permits unrestricted use, distribution, and reproduction in any medium for non-commercial purposes, provided the original author and source are credited, a link to the CC License is provided, and changes – if any – are indicated. First published online: September 11, 2017
ISSN 2062-8633 © 2017 The Author(s)
cobas® CT/NG for use on the cobas® 6800/8800 Systems
the United States of America (USA) and Australia have highlighted the importance of testing oropharyngeal and anorectal specimens in addition to urogenital tests in men who have sex with men (MSM) [9–14]. One study in San Francisco showed that 53% of C. trachomatis and 64% of N. gonorrhoeae infections in MSM involved non-urethral sites and would be missed if screening was done only for urethral infection [9]. In addition, the prevalence of anorectal CT in women attending STI clinics has been shown to be similar to that in MSM (up to 15% in women vs. 14% in MSM) [15–17]. Furthermore, recent studies in the USA looking at STI rates after the implementation of HIV preexposure prophylaxis (PrEP) with antiretroviral therapy have shown that no new HIV infections were noted with increasing use of HIV PrEP, despite high rates of STIs and decreased condom use. Thus, as PrEP use expands, STI screening will play an important role in patient management [18]. The diagnosis of infection with CT and NG is established with sensitive and specific NAATs. There are many commercially available choices for STI NAATs. The cobas® 4800 CT/NG Test addresses the needs for low to midthroughput testing for CT/NG [19]. The performance of the test has been confirmed in large clinical trials [20–22]. In many countries, centralized testing in large reference laboratories using high-throughput, automated NAAT systems has been the trend for large STI screening programs [23–25]. Therefore, cobas® CT/NG for use on the cobas® 6800/8800 Systems was recently launched addressing the needs of high-throughput laboratories with up to 960 samples per 8-h shift. In the present study, the analytical and clinical performance of cobas® CT/NG for use on the cobas® 6800/8800 Systems was demonstrated using a variety of urogenital, oropharyngeal, and anorectal specimens from female and male individuals.
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Methods cobas® CT/NG for use on the cobas® 6800 and cobas® 8800 Systems cobas® CT/NG is a qualitative nucleic acid test performed on the cobas® 6800 System and cobas® 8800 System. The test received CE-IVD mark in 2016. The system features, e.g., the instrument’s high-throughput capacity (384 samples per 8-h shift for the cobas® 6800 System and 960 samples per 8-h shift on the cobas® 8800 System), have recently been reviewed in detail [23]. Figure 1 shows the key features of the cobas® 6800 System and cobas® 8800 System. cobas® CT/NG enables the detection of CT/NG DNA from vaginal (clinician collected and self-collected with clinician instruction) and endocervical swabs, and female and male urine, as well as detection from female and male oropharyngeal and anorectal swabs collected and stabilized with cobas® PCR media. Cervical specimens collected in PreservCyt® solution were also evaluated. The volumes of sample required for the test are 400 μl for all swab samples and for PreservCyt® samples, and 850 μl for urine samples. Test duration is 37.5, suggesting target titers below the sensitivity of Sanger sequencing. Sequencing results for all discrepant NG specimens are shown in Table 4. Of the urogenital specimens, 8 of 12 NG discordant samples were confirmed by sequencing. Twenty of 22 discrepant oropharyngeal specimens (NG positive on the cobas® 6800/8800 Systems but negative on the cobas® 4800 System) were confirmed NG positive by Sanger sequencing, and all 8 anorectal specimens that were NG discrepant (cobas® 6800/8800 Test positive) were confirmed NG positive. In addition, 71 of 74 concordant NG positive oropharyngeal specimens were confirmed positive for NG by Sanger sequencing; similarly, 64 of 71 concordant NG positive anorectal specimens were confirmed NG positive by Sanger sequencing. The majority of PCR positive/ sequencing negative results showed Ct values at or below the sensitivity of Sanger sequencing.
Discussion Results presented in this study demonstrate the excellent performance of the cobas® 6800/8800 Systems for highthroughput CT/NG testing. Using these systems, centralized laboratories are able to handle mid to high volume CT/NG testing using both a large variety of established urogenital specimens as well as extragenital specimens (oropharyngeal and anorectal swabs). cobas® CT/NG for use on the cobas® 6800/8800 Systems is the first CE-IVD test that has been validated for use with extragenital specimens in addition to urogenital specimens. Studies have demonstrated the importance of testing extragenital specimens in certain high-risk populations and that >50% of infections may be missed in MSMs when only testing with traditional urogenital specimens [9–14]. In this regard, a recent study in a cohort of female patients attending a Dutch STD clinic revealed that routine universal anorectal screening for CT in women who do not report anal sex or symptoms is feasible, highly acceptable, and superior to the current selective testing on indication since 80% of anorectal CT infections would not have been tested for [17]. In contrast, selective testing appeared to be an appropriate control strategy for anorectal NG infection in women. The excellent analytical performance of cobas® CT/NG for use on the cobas® 6800/8800 Systems thus provides vastly improved laboratory efficiency for STI screening programs, i.e., all sample types seen in routine practice are validated and demonstrate excellent performance. Improved laboratory efficiencies may also be driven by increased screening in MSMs using PrEP. A recent metaanalysis reported that MSM using PrEP were 25.3 times more likely to acquire NG infection, 11.2 times more likely to acquire CT infection, and 44.6 times more likely to acquire syphilis versus MSM not using PrEP [26]. However,
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using a modeling approach, Jenness et al. [27] predicted an overall decrease in CT and NG infections in MSMs using PrEP based on adherence to CDC guidelines for MSMs using PrEP [28] with more frequent STD screening despite reduced condom use [27]. Analytical sensitivity of the cobas® 4800 CT/NG Test was found to be around 40 EB/ml for CT and 1 CFU/ml for NG in most specimen types and was as low as 20 EB/ml and 0.5 CFU/ml for urine samples. Clinical sensitivity expressed as percent agreement with the cobas® 4800 CT/NG Test was very high ranging from a PPA of ≥98.4% for CT and ≥95% for NG in all specimen types to an NPA and OPA of ≥98.6% for both CT and NG in all specimen types. Sanger sequencing confirmed the validity of the vast majority of 6800/8800 positive and 4800 negative results, thus demonstrating the superior sensitivity of cobas® CT/NG for use on the cobas® 6800/8800 Systems. Such high sensitivity is desirable as many STIs are asymptomatic but can readily be transmitted to partners. Importantly, cross-reactivity was not observed with other non-gonorrheal Neisseria strains. When testing mucosal sites, i.e., genital and especially oropharyngeal specimens, high specificity for NG and lack of cross-reactivity with non-gonorrheal Neisseria spp. is critical [29, 30]. In this regard, during development of cobas® CT/NG for use on the cobas® 6800/8800 Systems, algorithm parameters were optimized to decrease the likelihood of reporting a false-positive result. In the present study, exclusivity studies with a large number of non-gonorrheal Neisseria strains as well as results of discordant analysis using sequencing with a validated and distinct target region for clinical specimens confirmed the absence of cross-reactivity; this finding is further strengthened by the lack of cross-reactivity observed when sequencing extragenital samples that gave concordant results. Lastly, cross-contamination as determined using very high titer samples was very low (0.5%). In the screening population, we determined a theoretical cross-contamination rate of approximately 0.025%. This rate is noteworthy in light of the high-throughput capabilities of the systems allowing approximately 1000 samples to be run in an 8-h shift (see below and Fig. 1). The value of supporting large screening programs has been demonstrated through the identification of infected individuals and the initiation of treatment for the individual and partner(s). Altogether, these measures will support public health efforts to reduce the incidence of CT and NG infections [8]. Correlation analysis between cobas® CT/NG for use on the cobas® 6800/8800 Systems and the cobas® 4800 CT/NG Test found that the overall percent agreement was >95% for both CT and NG in all specimen types. Discordant analysis further demonstrated the improved sensitivity of cobas® CT/NG for use on the cobas® 6800/8800 Systems compared to the cobas® 4800 CT/NG Test at continued high specificity. For the PCR+/sequencing− results, the majority of Ct values were ≥37.5, suggesting target titers below the sensitivity of Sanger sequencing. The increased sensitivity of cobas® CT/NG for use on the cobas® European Journal of Microbiology and Immunology
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6800/8800 Systems is likely caused by changes that have been introduced to the assay. While primer and probe design are the same for the assays for use on the cobas® 4800 and the cobas® 6800/8800 Systems, continuous mixing of the sample during the lysis incubation step was introduced and allows for a higher rate of DNA recovery. In addition, a lower elution volume provides a more concentrated extract, also contributing to higher sensitivity. cobas® CT/NG for use on the cobas® 6800/8800 Systems offers simplicity and flexibility for mid to high volume laboratories. The cobas® 6800/8800 Systems currently offer the highest throughput solution for molecular CT/NG testing, achieving up to 384 CT/NG results in an 8-h shift on the cobas® 6800 System and up to 960 CT/NG results in an 8-h shift on the cobas® 8800 System. In addition, based on the large onboard storage capacity, enough CT/NG reagents, and positive and negative controls can be loaded to process more than 4600 results. With this large reagent capacity and an open stability for reagent cassettes of 90 days, the system offers a true walk-away solution. The operator interaction required for performance of the test is minimal and consists of loading of samples and consumables (at the start of a shift and as needed during shifts, depending on test mix and volume), clearing the waste containers from time to time, and managing the assays via onboard touch screen interface. Unlike other solutions for CT/NG testing, cobas® CT/NG offers ready to load reagent cartridges, with no reagent reconstitution, no pouring, no mixing, no thawing required. With a walk-away time of 8 h (cobas® 6800) or 4 h (cobas® 8800), staff can efficiently be used to perform other duties while samples are being processed. In a German laboratory performing high-throughput screening for CT [31], several mid-throughput platforms were retired after the introduction of a single cobas® 8800 System. Figure 2 shows performance characteristics for the cobas® 4800, cobas® 6800, and cobas® 8800 systems when run in batch mode (1 batch = 94 samples) at this site. The implementation of the high-throughput system allowed for an increased number of batches per day to be performed with a theoretical maximum of 16 batches
(1504 samples) in one working day (two shifts, total working hours 7 AM to 6.30 PM). At this time point, due to the very recent implementation of the cobas® 8800, only up to 12 batches are performed in a single working day. Thus, one cobas® 8800 alone can handle the same number of batches as five cobas® 4800 systems during the aforementioned laboratory hours; an even higher throughput could be achieved using the walk-away ability of the cobas® 8800 system, i.e., initiating runs at the end of the working day and releasing results the next morning. While the cobas® 6800/8800 Systems allow for test consolidation, it also allows for capacity to accommodate growing testing needs. For example, high-throughput testing is a hallmark of screening for cervical cancer. cobas® CT/NG for use on the cobas® 6800/8800 Systems has also been validated for detection of CT/NG in cervical specimens collected in PreservCyt® solution (commonly performed for cervical cancer screening via molecular tests and/or Pap smears). On the cobas® 6800 and 8800 Systems, CT/NG and HPV can be processed from the same cervical sample (aliquot from PreservCyt® solution) during the same run. In addition, all sample types for cobas® CT/NG can be processed during the same run as samples being processed using cobas® HPV. Furthermore, as testing menus expand on the cobas® 6800 and 8800 Systems, the ability to consolidate testing will become a key to a laboratories workflow and testing expansion. The cobas® 6800 and 8800 Systems are able to leverage common reagents, consumables, and processing profiles, where technically feasible. Using this approach will allow laboratories to process CT/NG and TV/MG (currently in development) from one patient sample during the same run with no sorting or reloading of patient samples, reducing the turn-around-time for delivering results back to a clinician. Since CT/NG and TV/MG are both combination tests, there would be enough patient sample available for cases where a repeat test or additional testing is required. Other reproductive health tests (i.e., Ureaplasma urealyticum) could be developed and run as laboratory-developed tests (LDTs) using the cobas® omni utility channel on the cobas®
Fig. 2. Comparison of system performances between the cobas® 4800, cobas® 6800, and cobas® 8800 Systems (1 run = 96 results, including controls) at a high-throughput customer site European Journal of Microbiology and Immunology
cobas® CT/NG for use on the cobas® 6800/8800 Systems
6800/8800 Systems. The cobas® omni utility channel allows laboratories to develop and run LDTs on the same system (processed during different runs) as their IVDs reducing instrument footprint and staff training requirements in the laboratory. The flexibility and ability to accommodate additional testing can provide laboratories with a system that allows for growth, efficiency, and administrative planning.
Conclusion cobas® CT/NG for use on the cobas® 6800 and 8800 Systems exhibited high sensitivity, specificity, PPV, and NPV for the detection of CT and NG in urogenital (endocervical swabs, urine specimens, and vaginal swabs collected in cobas® PCR Media and cervical specimens collected in PreservCyt® Solution) as well as extragenital (oropharyngeal and anorectal swabs) specimens to support screening of at risk populations. The unprecedented capacity for high-throughput testing of up to 960 samples in an 8-h shift makes the cobas® 6800 and 8800 Systems an attractive platform for molecular laboratories.
Funding sources This study was funded by Roche Molecular Systems, Pleasanton, CA.
Conflict of interest E.M., D.H., M.K., R.A., T.S., B.S., and O.L. are employees of Roche Molecular Diagnostics.
Acknowledgements We are grateful to Pari Hemyari, Enrique Marino, and Jesse Canchola for expert statistical support. We are also grateful to Kate Suileabhain and Michael Lewinski for critical review of the article.
References 1. Detels R, Green AM, Klausner JD, Katzenstein D, Gaydos C, Handsfield H, Pequegnat W, Mayer K, Hartwell TD, Quinn TC: The incidence and correlates of symptomatic and asymptomatic Chlamydia trachomatis and Neisseria gonorrhoeae infections in selected populations in five countries. Sex Transm Dis 38, 503–509 (2011) 2. Newman L, Rowley J, Vander Hoorn S, Wijesooriya NS, Unemo M, Low N, Stevens G, Gottlieb S, Kiarie J, Temmerman M: Global estimates of the prevalence and incidence of four curable sexually transmitted infections in 2012 based on systematic review and global reporting. PLoS One 10, e0143304 (2015)
185
3. Price MJ, Ades AE, De Angelis D, Welton NJ, Macleod J, Soldan K, Simms I, Turner K, Horner PJ: Risk of pelvic inflammatory disease following Chlamydia trachomatis infection: analysis of prospective studies with a multistate model. Am J Epidemiol 178, 484–492 (2013) 4. Gottlieb SL, Xu F, Brunham RC: Screening and treating Chlamydia trachomatis genital infection to prevent pelvic inflammatory disease: interpretation of findings from randomized controlled trials. Sex Transm Dis 40, 97–102 (2013) 5. Rekart ML, Gilbert M, Meza R, Kim PH, Chang M, Money DM, Brunham RC: Chlamydia public health programs and the epidemiology of pelvic inflammatory disease and ectopic pregnancy. J Infect Dis 207, 30–38 (2013) 6. Romoren M, Hussein F, Steen TW, Velauthapillai M, Sundby J, Hjortdahl P, Kristiansen IS: Costs and health consequences of chlamydia management strategies among pregnant women in sub-Saharan Africa. Sex Transm Infect 83, 558–566 (2007) 7. Sexton J, Garnett G, Rottingen JA: Metaanalysis and metaregression in interpreting study variability in the impact of sexually transmitted diseases on susceptibility to HIV infection. Sex Transm Dis 32, 351–357 (2005) 8. Workowski KA, Bolan GA, Centers for Disease C, Prevention: Sexually transmitted diseases treatment guidelines, 2015. MMWR Recomm Rep 64, 1–137 (2015) 9. Kent CK, Chaw JK, Wong W, Liska S, Gibson S, Hubbard G, Klausner JD: Prevalence of rectal, urethral, and pharyngeal chlamydia and gonorrhea detected in 2 clinical settings among men who have sex with men: San Francisco, California, 2003. Clin Infect Dis 41, 67–74 (2005) 10. Annan NT, Sullivan AK, Nori A, Naydenova P, Alexander S, McKenna A, Azadian B, Mandalia S, Rossi M, Ward H, Nwokolo N: Rectal chlamydia – a reservoir of undiagnosed infection in men who have sex with men. Sex Transm Infect 85, 176–179 (2009) 11. Gunn RA, O’Brien CJ, Lee MA, Gilchick RA: Gonorrhea screening among men who have sex with men: value of multiple anatomic site testing, San Diego, California, 1997–2003. Sex Transm Dis 35, 845–848 (2008) 12. Ota KV, Fisman DN, Tamari IE, Smieja M, Ng LK, Jones KE, Diprima A, Richardson SE: Incidence and treatment outcomes of pharyngeal Neisseria gonorrhoeae and Chlamydia trachomatis infections in men who have sex with men: a 13-year retrospective cohort study. Clin Infect Dis 48, 1237–1243 (2009) 13. Schachter J, Moncada J, Liska S, Shayevich C, Klausner JD: Nucleic acid amplification tests in the diagnosis of chlamydial and gonococcal infections of the oropharynx and rectum in men who have sex with men. Sex Transm Dis 35, 637–642 (2008) 14. Centers for Disease Control and Prevention: Recommendations for the laboratory-based detection of Chlamydia trachomatis and Neisseria gonorrhoeae – 2014. MMWR Recomm Rep 63, 1–19 (2014) 15. Javanbakht M, Gorbach P, Stirland A, Chien M, Kerndt P, Guerry S: Prevalence and correlates of rectal chlamydia and gonorrhea among female clients at sexually transmitted disease clinics. Sex Transm Dis 39, 917–922 (2012) 16. van Liere GA, Hoebe CJ, Wolffs PF, Dukers-Muijrers NH: High co-occurrence of anorectal chlamydia with urogenital chlamydia in women visiting an STI clinic revealed by routine universal testing in an observational study; a recEuropean Journal of Microbiology and Immunology
186
17.
18.
19.
20.
21.
22.
23.
E. M. Marlowe et al.
ommendation towards a better anorectal chlamydia control in women. BMC Infect Dis 14, 274 (2014) van Liere G, Dukers-Muijrers N, Levels L, Hoebe C: High proportion of anorectal Chlamydia trachomatis and Neisseria gonorrhoeae after routine universal urogenital and anorectal screening in women visiting the sexually transmitted infection clinic. Clin Infect Dis 64, 1705–1710 (2017) Volk JE, Marcus JL, Phengrasamy T, Blechinger D, Nguyen DP, Follansbee S, Hare CB: No new HIV infections with increasing use of HIV preexposure prophylaxis in a clinical practice setting. Clin Infect Dis 61, 1601–1603 (2015) Van Der Pol B: Cobas(R) 4800: a fully automated system for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae. Expert Rev Mol Diagn 13, 131–140 (2013) Van Der Pol B, Liesenfeld O, Williams JA, Taylor SN, Lillis RA, Body BA, Nye M, Eisenhut C, Hook EW 3rd: Performance of the cobas CT/NG test compared to the Aptima AC2 and Viper CTQ/GCQ assays for detection of Chlamydia trachomatis and Neisseria gonorrhoeae. J Clin Microbiol 50, 2244–2249 (2012) Van Der Pol B, Taylor SN, Liesenfeld O, Williams JA, Hook EW, 3rd: Vaginal swabs are the optimal specimen for detection of genital Chlamydia trachomatis or Neisseria gonorrhoeae using the Cobas 4800 CT/NG test. Sex Transm Dis 40, 247–250 (2013) Taylor SN, Liesenfeld O, Lillis RA, Body BA, Nye M, Williams J, Eisenhut C, Hook EW 3rd, Van Der Pol B: Evaluation of the Roche cobas(R) CT/NG test for detection of Chlamydia trachomatis and Neisseria gonorrhoeae in male urine. Sex Transm Dis 39, 543–549 (2012) Cobb B, Simon CO, Stramer SL, Body B, Mitchell PS, Reisch N, Stevens W, Carmona S, Katz L, Will S, Liesenfeld O: The cobas(R) 6800/8800 System: a new era of auto-
European Journal of Microbiology and Immunology
24.
25.
26.
27.
28.
29.
30.
31.
mation in molecular diagnostics. Expert Rev Mol Diagn 17, 167–180 (2017) Zaninotto M, Plebani M: The “hospital central laboratory”: automation, integration and clinical usefulness. Clin Chem Lab Med 48, 911–917 (2010) Ledeboer NA, Dallas SD: The automated clinical microbiology laboratory: fact or fantasy? J Clin Microbiol 52, 3140–3146 (2014) Kojima N, Davey DJ, Klausner JD: Pre-exposure prophylaxis for HIV infection and new sexually transmitted infections among men who have sex with men. AIDS 30, 2251– 2252 (2016) Jenness SM, Weiss KM, Goodreau SM, Gift T, Chesson H, Hoover KW, Smith DK, Liu AY, Sullivan PS, Rosenberg ES: Incidence of gonorrhea and chlamydia following HIV preexposure prophylaxis among men who have sex with men: a modeling study. Clin Infect Dis 65, 712–718 (2017), DOI: 10.1093/cid/cix439 (2017) US Public Health Service Preexposure prophylaxis for the prevention of HIV infection in the United States – 2014: a clinical practice guideline. https://wwwcdcgov/hiv/pdf/ prepguidelines2014pdf (2014) Tabrizi SN, Unemo M, Limnios AE, Hogan TR, Hjelmevoll SO, Garland SM, Tapsall J: Evaluation of six commercial nucleic acid amplification tests for detection of Neisseria gonorrhoeae and other Neisseria species. J Clin Microbiol 49, 3610–3615 (2011) Upton A, Bromhead C, Whiley DM: Neisseria gonorrhoeae false-positive result obtained from a pharyngeal swab by using the Roche cobas 4800 CT/NG assay in New Zealand in 2012. J Clin Microbiol 51, 1609–1610 (2013) Bohm I, Groning A, Sommer B, Muller HW, Krawczak M, Glaubitz R: A German Chlamydia trachomatis screening program employing semi-automated real-time PCR: results and perspectives. J Clin Virol 46 (Suppl 3), S27–S32 (2009)