Performance of Nucleic Acid Amplification Tests for the Detection of Neisseria Gonorrhoeae and Chlamydia Trachomatis in Extragenital Sites
Phase: NA
Conditions: Neisseria Gonorrhoeae Infection, Chlamydia Trachomatis Infection, Sexually Transmitted Infections
Sponsor: Duke University

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Antibacterial Resistance Leadership Group (ARLG) 
Performance of Nucleic Acid Amplification Tests for the Detection of Neisseria gonorrhoeae 
and Chlamydia trachomatis in Extragenital Sites  
(pNAAT) 
Protocol Number:  
ARLG_pNAAT-Yr3 
Funding Sponsor: 
National Institute of Allergy and Infectious Diseases (NIAID) 
Funding Mechanism: 5UM1AI104681 
Protocol Date: 
December 4th, 2017 
Protocol Version: 
4.0 
Principal Investigator 
Dr. Jeffrey D. Klausner, MD, MPH 
NCT02870101

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Signature Page 
 
 
The signature below documents the review and approval of this protocol and provides the necessary 
assurances that this study will be conducted according to the protocol, including all statements regarding 
confidentiality, and according to national, regional, and local legal and regulatory requirements. 
 
 
   
 
 
Site Principal Investigator Name (Print)  
 
 
 
 
 
 
Signature 
 
Date 
 
 
 
 
 

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ARLG GC Protocol Team Roster 
All questions concerning this protocol should be sent via email to arlg.gc@mc.duke.edu.  The appropriate 
team member will respond to questions via email, generally within 24 hours (Monday-Friday).
Protocol Chair 
 
Jeffrey D. Klausner, MD, MPH 
Professor of Medicine and Public Health 
University of California, Los Angeles 
10833 Le Conte Avenue 
Los Angeles, CA 90095 
Phone: 310-267-0409 
Email: JDKlausner@mednet.ucla.edu 
 
Project Leader 
 
Zoe Sund 
Duke Clinical Research Institute 
2400 Pratt Street 
Durham NC 27705 
Phone:  919-428-1007 
Email:  zoe.sund@duke.edu  
 
Statisticians 
 
Scott Evans, PhD 
Harvard School of Public Health FXB 513 
651 Huntington Avenue 
Boston, MA 02115-6017 
Phone: 617-432-2998 
Email: evans@sdac.harvard.edu 
 
Lauren Komarow, MSc 
Harvard School of Public Health FXB 549A 
651 Huntington Avenue 
Boston, MA 02115-6017 
Phone: 617-432-3233 
Email: lkomarow@sdac.harvard.edu  
 
Thuy Tran, MSc 
Harvard School of Public Health FXB 608 
651 Huntington Avenue 
Boston, MA 02115-6017 
Phone: 617-432-7523 
Email: ttran@sdac.harvard.edu 
 
 
 
 
Protocol Clinicians 
 
Sarah Doernberg, MD, MAS 
Division of Infectious Diseases  
University of California, San Francisco 
513 Parnassus Ave, Box 0654 
San Francisco, CA 94143  
Phone: 415-502-5548 
Email: sarah.doernberg@ucsf.edu 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 

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Table of Contents 
LIST OF ABBREVIATIONS AND DEFINITIONS ............................................................................................. 6 
1.0 
BACKGROUND AND SCIENTIFIC RATIONALE .............................................................................. 9 
1.1 BACKGROUND INFORMATION ................................................................................................................... 9 
1.2 SCIENTIFIC RATIONALE ............................................................................................................................ 9 
2.0 
OBJECTIVES ....................................................................................................................................... 11 
2.1 PRIMARY OBJECTIVE ............................................................................................................................. 11 
2.2 SECONDARY OBJECTIVES ...................................................................................................................... 11 
2.3 OUTCOME MEASURES ........................................................................................................................... 11 
3.0 
STUDY DESIGN .................................................................................................................................. 14 
3.1 STUDY DESIGN ....................................................................................................................................... 14 
3.2 INCLUSION CRITERIA .............................................................................................................................. 15 
3.3 EXCLUSION CRITERIA ............................................................................................................................ 15 
4.0 
STUDY PROCEDURES ...................................................................................................................... 16 
4.1 RECRUITMENT PLAN .............................................................................................................................. 16 
4.2 SCREENING ............................................................................................................................................ 16 
4.3 ENROLLMENT ......................................................................................................................................... 16 
4.6 FOLLOW-UP VISIT ................................................................................................................................... 17 
5.0 
DEVICE MONITORING ....................................................................................................................... 17 
5.1 SPECIMEN COLLECTION DEVICE MONITORING ..................................................................................... 17 
5.2 LABORATORY DEVICE MONITORING ...................................................................................................... 17 
6.0 
STATISTICS......................................................................................................................................... 18 
6.1 STUDY DESIGN ....................................................................................................................................... 18 
6.2 ENDPOINTS ............................................................................................................................................ 18 
6.3 SAMPLE SIZE .......................................................................................................................................... 18 
6.4 RANDOMIZATION OF SWABS .................................................................................................................. 19 
6.5 MONITORING .......................................................................................................................................... 19 
6.6 ANALYSES .............................................................................................................................................. 20 
7.0 
QUALITY CONTROL .......................................................................................................................... 23 
7.1 QUALITY CONTROLS AND STUDY MONITORING .................................................................................... 23 
7.2 SOURCE DOCUMENTS AND ACCESS TO SOURCE DATA ....................................................................... 23 
8.0 
ETHICS / PROTECTION OF HUMAN SUBJECTS .......................................................................... 23 
8.1 ETHICAL STANDARDS ............................................................................................................................. 23 
8.2 INSTITUTIONAL REVIEW BOARD AND INFORMED CONSENT .................................................................. 23 
8.3 SUBJECT CONFIDENTIALITY ................................................................................................................... 23 
8.4 SPECIMEN HANDLING ............................................................................................................................. 24 

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9.0 
DATA HANDLING AND RECORD KEEPING .................................................................................. 24 
9.1 DATA COLLECTION ................................................................................................................................. 24 
9.2 INSPECTION OF RECORDS ..................................................................................................................... 24 
9.3 RETENTION OF RECORDS ...................................................................................................................... 24 
9.4 CONFIDENTIALITY ................................................................................................................................... 25 
10.0 
PUBLICATION POLICY ...................................................................................................................... 25 
APPENDIX A - SCHEDULE OF EVENTS ...................................................................................................... 30 

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List of Abbreviations and Definitions 
ARLG 
Antibacterial Resistance Leadership Group 
ASIS 
Anatomic Site Infection Status 
CDC 
Center for Disease Control and Prevention 
CFR 
Code of Federal Regulations 
CT 
Chlamydia trachomatis 
DCRI 
Duke Clinical Research Institute 
DNA 
Deoxyribonucleic acid 
Equivocal 
Final equivocal result from nucleic acid amplification test, as determined by 
manufacturer’s guidance. This means the test was run and the result was neither 
positive nor negative 
FDA 
Food and Drug Administration 
HIV 
Human immunodeficiency virus 
ICMJE 
International Committee of Medical Journal Editors 
ID 
Identification 
IFU 
Instructions for Use 
Indeterminate 
Anatomic site infected standard cannot be determined as positive or negative 
IRB 
Institutional Review Board 
ISRC 
Independent Study Review Committee 
LGBT 
Lesbian, gay, bisexual and transgender 
NAAT 
Nucleic acid amplification test 
Negative (-) 
NAAT reading 
Result from the nucleic acid amplification test is negative 
NR 
This means no test was run and there is no result 
NPA 
Negative percent agreement 
NPV 
Negative predictive value 
NG 
Neisseria gonorrhoeae 
NIH 
National Institutes of Health 
OHRP 
Office of Human Research Protections 
Positive (+) 
NAAT reading 
Result from the nucleic acid amplification test is positive 
PPA 
Positive percent agreement 
PPV 
Positive predictive value 
PI 
Principal Investigator 
RNA 
Ribonucleic acid 

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rRNA 
Ribosomal ribonucleic acid 
SDMC 
Statistical and Data Monitoring Center 
STD 
Sexually transmitted diseases 
WHO 
World Health Organization 

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Protocol Synopsis 
Protocol Title: 
Performance of Nucleic Acid Amplification Tests for the Detection of 
Neisseria gonorrhoeae and Chlamydia trachomatis in Extragenital Sites 
Study Design: 
A cross-sectional, single visit study to evaluate the diagnostic accuracy of 
nucleic acid amplification tests for detection of Neisseria gonorrhoeae and 
Chlamydia trachomatis from pharyngeal and rectal sites 
Primary Study 
Objectives: 
For each nucleic acid amplification assay, estimate the positive percent 
agreement (PPA) and negative percent agreement (NPA) for detecting: 
•
Neisseria gonorrhoeae in rectal swabs
•
Neisseria gonorrhoeae in pharyngeal swabs
•
Chlamydia trachomatis in rectal swabs
•
Chlamydia trachomatis in pharyngeal swabs
Study Population 
Patients presenting to an outpatient clinic for sexually transmitted disease 
(STD) testing for Neisseria gonorrhoeae and Chlamydia trachomatis 
infections 
Number of 
subjects 
Up to 3,000 
Study duration 
Approximately 12 to 24 months 
Number of sites 
Up to 10 clinical sites (as needed) 
Clinical Samples 
Pharyngeal and rectal swabs 

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1.0 
Background and Scientific Rationale 
1.1 
Background Information 
Nucleic acid amplification tests (NAATs) have become the gold standard for diagnosing 
Neisseria gonorrhoeae (NG) and Chlamydia trachomatis (CT) infections in the urogenital 
tract but are not currently approved by the Food and Drug Administration (FDA) for use in 
extragenital sites, including the pharynx and the rectum.  
The goal of this study is to determine the diagnostic accuracy of the following 3 NAAT 
platforms for the detection of NG and CT from the pharynx and the rectum:  
•
Company 1 Assay [Company 1] 1
•
Company 2 Assay [Company 2] 2
•
Company 3 Assay [Company 3] 3
The assays being evaluated are intended for the direct qualitative detection of Chlamydia 
trachomatis and/or Neisseria gonorrhoeae in rectal or pharyngeal clinician-collected 
swabs from symptomatic or asymptomatic male or female patients. 
1.2 
Scientific Rationale 
Infections due to NG and CT are a major public health threat. The World Health 
Organization (WHO) estimated more than 100 million new worldwide cases for each of 
NG and CT in 2008.4  In the US alone, there were 333,000 NG and 1.4 million CT 
genitourinary infections in the year 2013.5 Over the past 15 years, researchers have 
demonstrated an important burden of NG and CT infection in extragenital sites - the 
pharynx and rectum. Most CT and NG infections are asymptomatic, but infection can lead 
to serious sequelae, including infertility, chronic pelvic pain, adverse obstetrical 
outcomes, increased risk of human immunodeficiency virus (HIV) acquisition, and 
disseminated infection.6-9 In addition to threats to individual health, there has been 
growing concern over antibiotic resistance, and in 2013, the US Centers for Disease 
Control and Prevention (CDC) classified drug-resistant NG as one of the three urgent-
level resistant bacteria. Improved detection of extragenital NG is thought to be a crucial 
component of adequate treatment and thus prevention of further resistance.10  
Currently, the CDC recommends the use of NAATs for NG and CT screening and 
diagnosis in the genitourinary tract due to superior sensitivity compared to traditional 
culture methods.11 The sensitivity and specificity of commercially available NAATs for the 
detection of genitourinary NG and CT infection is estimated to be between 90-100%.12  
CDC also recommends the use of NAATs for extragenital sites.11 Due to widespread use 
and interest in these tests for extragenital NG and CT infections, multiple studies have 
examined the test characteristics of NAAT tests in comparison to culture and to additional 
NAAT platforms (see Table 1). For diagnosis of CT, reported sensitivities are 80-100% 
(pharyngeal) and 46-100% (rectal), with significant variation by platform tested.13-19 
Reported specificities are >99% for pharyngeal site and 89-100% for the rectum. For 
diagnosis of NG, reported sensitivities are 72-100% (pharyngeal) and 75-100% (rectal), 
again with variation by platform.13,14,16-18,20,21 Reported specificities are 72-100% 
(pharyngeal) and 95-100% (rectal).  

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Table 1. Prior Studies of extragenital testing for chlamydia and gonorrhea. 
Screening test 
Sensitivity/specificity* by specimen type (%) 
Pharyngeal 
Rectal 
Neisseria gonorrhoeae 
 Bachmann LH et al, 201016 
100 / 96.2 
100 / 95.5 
 Cosentino LA et al, 201217 
76 / 100 
 Moncada J et al, 200919 
78.3 / 99.8 
 Ota KV et al, 200913 
95 / 99.6 
100 / 100 
 Schachter J et al, 200814 
84.3 / 99.4 
93.2 /  99.7 
100/100 
100/100 
Walsh A et al, 201122 
 Goldenberg SD et al., 201223 
91.1 / 100 
 Cosentino LA et al, 201524 
100 / 100 
Chlamydia trachomatis 
 Bachmann LH et al, 201016 
100 / 88.8 
 Cosentino LA et al, 201217 
100 / 99.8 
 Moncada J et al, 200919 
71.2 / 99 
 Ota KV et al, 200913 
100 / 99.2 
100 / 98.7 
 Schachter J et al, 200814 
100 / 99.6 
93.5 / 97.7 
 Goldenberg SD et al., 201223 
86 / 99.2 
 Cosentino LA et al, 201524 
96.6 / 99.7 
*Sensitivity and specificity as reported by authors, each using unique definitions for the reference standard. As
discussed below, we will be using positive percent agreement (PPA) and negative percent agreement (NPA) for
these calculations.
Despite the CDC recommendations, at this time, there are no FDA-cleared commercial 
NAAT tests for the detection of pharyngeal and rectal NG or CT infection.  The FDA-
clearance of such assays would increase the availability and uptake and subsequently 
lead to improved screening and enhanced control of NG and CT extragenital infections.  
The current proposed study will help provide the data required for FDA consideration to 
clear such assays for marketing. 
Because of the multi-site nature of the ARLG, its strong laboratory network and inclusion 
of investigators with expertise in NG and CT detection and treatment, the ARLG is well-
positioned to conduct such a study.  Current commercial assays are “dual” assays and 
include targets for both NG and CT, so the goal of this study will be to evaluate the 
diagnostic accuracy for detection of both organisms at extragenital sites.   
The molecular targets of the three NAAT platforms that will be tested differ, which will 
allow for comparative evaluation.25  
•
Company 1 Assay (Company 1) [redacted] 1

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•
Company 2 Assay (Company 2) [redacted] 2
•
Company 3 Assay (Company 3) [redacted] 3
Additional testing using the tiebreaker test [redacted] will be performed in cases of 
discordant results (see Section 3.5). This test utilizes [redacted] to identify the presence 
of the organism of interest in the clinical sample. [redacted]. Though the diagnostic 
accuracy of these assays for extragenital infections has not previously been evaluated, 
studies evaluating the performance of these assays for confirmation of other NAATs 
suggest excellent concordance.28-31 
2.0 
Objectives 
2.1 
Primary Objective 
For each NAAT, estimate the PPAs and NPAs for detecting: 
•
Neisseria gonorrhoeae in rectal swabs
•
Neisseria gonorrhoeae in pharyngeal swabs
•
Chlamydia trachomatis in rectal swabs
•
Chlamydia trachomatis in pharyngeal swabs
2.2 
Secondary Objectives 
2.2.1 
Global analyses: 
For each NAAT, positive predictive values (PPVs), negative predictive values (NPVs), 
positive likelihood ratios, negative likelihood ratios, and sensitivity analyses will be 
calculated for detecting:  
•
Neisseria gonorrhoeae in rectal swabs
•
Neisseria gonorrhoeae in pharyngeal swabs
•
Chlamydia trachomatis in rectal swabs
•
Chlamydia trachomatis in pharyngeal swabs
2.2.2 
Subgroup analyses: 
For each NAAT, to estimate the PPAs, NPAs, PPVs, and NPVs for detection of NG and 
CT from rectal and pharyngeal swab specimens by sex and by anatomic site-specific 
symptom status. 
2.3 
Outcome Measures 
The anatomic site infected status (ASIS) will be determined for each anatomic site 
(pharyngeal and rectal) and each organism (NG and CT), as shown below in Table 2. 

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Possible ASIS outcomes are: 
•
Infected
•
Not infected
•
Indeterminate
•
Invalid, exclude from analysis
The anatomic site is considered to be infected if both of the reference test results are 
positive. The anatomic site is considered to be not infected when both reference test 
results are negative. If there is discordance between the reference tests, an additional 
NAAT will be done as a tiebreaker. In this case, agreement of 2/3 of the reference NAATs 
will determine the ASIS. If two tests are equivocal or one equivocal and one not run, the 
third test result will stand as the ASIS if positive or negative. If two tests are not run, the 
ASIS will be considered invalid and will be excluded from the analysis. All possible 
combinations are shown in Table 2. The tiebreaker test will be run by the lab if any NAAT 
is not concordant with the others and interpreted only in the case of discordant results 
between the two planned reference tests for each assay.  [redacted]. 
To determine the ASIS, the test result for each respective site (pharyngeal or rectal) and 
each organism (NG or CT) for each NAAT platform will be used.  Possible outcomes for 
each assay are:  
•
Company 1 Assay (Company 1)1: Not detected, detected, invalid (sample 
processing control or sample adequacy control failed), error (probe check control 
failed), or no result (insufficient data was collected, e.g. test aborted). Initial invalid, 
error, or no result tests will be repeated. If the repeat test returns invalid, error, or no 
result, the final result will be considered an invalid and will be categorized as no result 
(NR) for the ASIS determination and statistical analyses below. If the repeat test 
returns not detected (negative) or detected (positive), this will be the result used for 
the ASIS determination and statistical analyses.
•
Company 2 Assay (Company 2)2: Negative, positive, equivocal (result between 
positive and negative), invalid (run status is FAIL or other technical failure), or error 
(sample was not tested due to an error detected by the instrument). Initial equivocal, 
invalid, and error test results will be repeated. If the repeat test result returns 
equivocal, the final test result will be considered an equivocal test result for the ASIS 
determination and statistical analyses below. If the repeat test result returns invalid or 
error, the final test result will be categorized as equivocal if the initial test was 
equivocal and as no result (NR) if the initial test was invalid or error for the ASIS 
determination and statistical analyses below. If the repeat test returns negative or 
positive, this will be the result considered for the ASIS determination and statistical 
analyses.
•
Company 3 Assay (Company 3)3: For NG, possible results are positive
(detected, with cycle number less than or equal to the assay cut-off), negative (no 
evidence of amplification or cycle number greater than the assay cut-off), and error. 
An equivocal interpretation does not apply. For CT, possible results are positive
(detected, with cycle number less than or equal to the assay cut-off), negative (no 
evidence of amplification), equivocal (cycle number beyond the assay cut-off), and 
error. A sample with initial interpretation of error (both CT and NG) or equivocal (CT 
only) will be retested. If the repeat test returns negative or positive, this will be the 
result considered for the ASIS determination and statistical analyses. If the repeat

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test result is equivocal (CT only), the final test result will be considered equivocal for 
the ASIS determination and statistical analyses below. If the repeat test result is 
error, the final test result will be categorized as no result (NR) for the ASIS 
determination and statistical analyses below if the initial test result was error and as 
equivocal if the initial test result was equivocal (CT only).  
•
Tiebreaker assay: Tiebreaker Assay (Company 4):  Negative, positive, equivocal 
(result between negative and positive ranges), invalid (run status is FAIL or other 
technical failure), or error (sample was not tested due to an error detected by the 
instrument). Initial equivocal, invalid and error test results will be repeated. If the 
repeated test result is equivocal, it will be considered an equivocal test result for the 
ASIS determination and statistical analyses below. If the repeat test result returns 
invalid or error, the final test result will be categorized as equivocal if the initial test 
was equivocal and as no result (NR) if the initial test was invalid or error for the ASIS 
determination and statistical analyses below. If the repeat test returns negative or 
positive, this will be the result considered for the ASIS determination and statistical 
analyses.
       Table 2. Determination of the Anatomic Site Infected Standard 
Comparator NAAT 1 
Result 
Comparator NAAT 2 
Result 
Tiebreaker NAAT 
Result 
Anatomic Site Infection 
status 
+ 
+ 
Not indicated 
Infected 
+ 
- 
+ 
Infected 
+ 
E* 
+ 
Infected 
+ 
NR** 
+ 
Infected 
+ 
- 
- 
Not infected 
+ 
- 
E 
Indeterminate 
+ 
- 
NR 
Indeterminate 
+ 
E 
- 
Indeterminate 
+ 
E 
E 
Infected 
+ 
E 
NR 
Infected 
+ 
NR 
- 
Indeterminate 
+ 
NR 
E 
Infected 
+ 
NR 
NR 
Invalid,  
remove from analysis 
- 
- 
Not indicated 
Not infected 
- 
+ 
- 
Not infected 
- 
E 
- 
Not infected 
- 
NR 
- 
Not infected 
- 
+ 
+ 
Infected 
- 
+ 
E 
Indeterminate 
- 
+ 
NR 
Indeterminate 
- 
E 
+ 
Indeterminate 
- 
E 
E 
Not infected 

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- 
E 
NR 
Not infected 
- 
NR 
+ 
Indeterminate 
- 
NR 
E 
Not infected 
- 
NR 
NR 
Invalid,  
remove from analysis 
E 
+ 
+ 
Infected 
E 
- 
- 
Not infected 
E 
+ 
- 
Indeterminate 
E 
+ 
E 
Infected 
E 
+ 
NR 
Infected 
E 
- 
+ 
Indeterminate 
E 
- 
E 
Not infected 
E 
- 
NR 
Not infected 
E 
NR 
+ 
Infected 
E 
NR 
- 
Not infected 
E 
NR 
E 
Indeterminate 
E 
NR 
NR 
Invalid,  
remove from analysis 
NR 
+ 
+ 
Infected 
NR 
- 
- 
Not infected 
NR 
NR 
Not indicated 
Invalid,  
remove from analysis 
NR 
+ 
- 
Indeterminate 
NR 
+ 
E 
Infected 
NR 
+ 
NR 
Invalid,  
remove from analysis 
NR 
- 
+ 
Indeterminate 
NR 
- 
E 
Not infected 
NR 
- 
NR 
Invalid,  
remove from analysis 
NR 
E 
+ 
Infected 
NR 
E 
- 
Not infected 
NR 
E 
E 
Indeterminate 
NR 
E 
NR 
Invalid,  
remove from analysis 
*E = equivocal result
**NR = no result. This can occur either because the test result was invalid or because the test could not be run
(e.g. too little sample, improperly shipped, no sample received).
3.0 
Study Design 
3.1 
Study Design 

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This will be a cross-sectional, single visit study evaluating the performance of three 
commercial NAATs to detect NG and CT in the rectum and pharynx.1,2,32,33  A fourth 
NAAT is being used as a tiebreaker.26,27 We are not evaluating the performance of this 
fourth NAAT.  
The study will include specimens collected from adult subjects seeking sexually 
transmitted disease (STD) testing at the participating clinics, which include clinics 
focused on sexually transmitted diseases, women’s health, student health, family 
planning clinics, and clinics specializing in lesbian, gay, bisexual, and transgender 
(LGBT) health.  Potential subjects will be identified, assessed for eligibility and 
approached for informed consent.  Both symptomatic and asymptomatic individuals will 
be included in the study population. 
The study will enroll up to 3,000 subjects who will have four pharyngeal and four rectal 
swabs collected as part of a one-time study visit.  Each manufacturer will have specific 
collection kit and transport media.  Additional data collection will include, but is not limited 
to, eligibility screening questions, gender, age, and patient report of relevant pharyngeal, 
rectal, or other relevant symptoms, and swab collection completeness. Subjects will be 
classified as symptomatic or asymptomatic for each anatomic site. Results of testing at 
other anatomic sites and results of clinical testing will not be available.  
Each swab will be used for a specific NAAT, which will be run at one of the two reference 
testing laboratories.  The ASIS will be defined by the results of the two other nucleic acid 
amplification tests, and a tiebreaker test (if necessary), as shown in Table 2. Each 
anatomic site will be considered in isolation.  
When accrual is complete, the study will be closed and an analysis plan will be enacted 
to finalize the database for analysis. 
3.2 
Inclusion Criteria 
a)
Attending a participating clinic for evaluation of STDs
b)
≥18 years of age at date of screening
c)
Able and willing to provide informed consent
d)
Willing to comply with study procedures, including collection of 4 swabs each
from the pharynx and rectum for NG and CT testing
NOTE:  Male, female, and transgender individuals will be included, and both symptomatic 
and asymptomatic individuals will be included. 
3.3 
Exclusion Criteria 
a)
Receipt of any systemic antibacterial drug in the past 14 days
b)
Receipt of myelosuppressive chemotherapy in the past 30 days
3.4 
Nucleic Acid Amplification Tests (NAATs) 
This study will evaluate three distinct NAAT platforms. The selected platforms25 are: 

Company 1 Assay (Company 1) [redacted] 1

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
Company 2 Assay (Company 2) [redacted].2
•
Company 3 Assay (Company 3) [redacted] 3
3.5 
Tiebreaker Assay 
In case of disagreement between reference NAAT results, a tiebreaker assay will be 
performed. This assay will use [redacted] (see Section 2.3). [redacted]   
4.0 
Study Procedures 
4.1 
Recruitment Plan 
This study will enroll up to 3,000 participants who are seeking STD testing at one of the 
participating clinics. Potential subjects will be recruited without regard for risk behavior, 
symptom status, or gender. Participating clinics may include those designed for STD 
screening and management, family planning, student health, women’s health, HIV 
management, and clinics focusing on the LGBT population.  
4.2 
Screening 
Participants will be recruited from clinical sites performing routine STD testing.  It is 
assumed that individuals presenting for routine STD testing at the participating clinics are 
at risk for such infections.  Each person will undergo screening for eligibility and, if 
eligible, will be approached for oral informed consent.   
4.3 
Enrollment 
Once enrolled, eight swabs will be taken: four swabs will be taken from the pharynx and 
four swabs will be taken from the rectum in addition to any swabs taken as part of routine 
clinical care. The swabs collected for routine clinical care will be taken first. The order of 
the research swabs will be randomized per subject to account for the possibility that the 
yield of the swab is affected by previous swabs at the same anatomic site. The swabs will 
be stored and transported per manufacturer guidelines. Results of those NAATs will not 
be reported back to the treating clinician for clinical purposes.  
Data collection will include, but is not limited to, assessment of signs and symptoms of 
rectal and/or pharyngeal infection, as well as socio-demographic information such as age 
(not date of birth), race, ethnicity and gender, and swab collection completeness.   
4.5 
Incomplete Swab Collection 

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While required as an inclusion criteria for a potential subject to agree to both pharyngeal 
and rectal swabs, if only one anatomic site is swabbed after enrollment for whatever 
reason, and all four swabs are completed for that anatomic site, those swabs for that 
anatomic site will be included in the analysis. If fewer than four swabs at a site are 
completed, that anatomic site (and swab results) will be excluded from the analysis. 
4.4 
Subject Withdrawal and Replacement 
Subjects may voluntarily withdraw their consent for study participation at any time and for 
any reason, without penalty.  The primary reason for withdrawal from the study will be 
recorded on the Study Status case report form.  Subjects who withdraw will not be 
replaced. 
4.5 
Study Discontinuation 
This study may be terminated at any time by the Principal Investigator (PI) in 
consultation with the Antibacterial Resistance Leadership Group (ARLG) and/or the 
National Institutes of Health (NIH).   
4.6 
Follow-up Visit 
There will not be a follow-up visit as part of this study. Subjects will continue with routine 
clinical care as directed by their medical providers. 
4.7 
Central Laboratory Procedures 
There will be two reference testing laboratories, which will divide the processing 
and running of the NAATs for the study. Each clinical study site will be assigned 
to send swabs to one of the two laboratories. Swabs will be processed and run at 
the reference testing laboratories according to each manufacturer’s Instructions 
for Use (IFU). Initial equivocal, invalid, or otherwise undetermined results will be 
repeated per manufacturer’s guidelines before determining whether the result will 
be classified as positive, negative, equivocal, or no result. One of the reference 
testing laboratories will be responsible for running all of the tiebreaker assays, 
regardless of the location of original study site. 
The reference testing laboratories will receive training for each of the study 
platforms from a representative from each of the manufacturers, in order to 
ensure adherence to the manufacturer’s IFU procedures.  
5.0 
Device Monitoring 
5.1 
Specimen Collection Device Monitoring 
Any complications with the specimen collection process, including device failure, 
significant patient discomfort, or inability to collect specimen, will be reported on the 
specimen collection case report form. 
5.2 
Laboratory Device Monitoring 
The reference testing laboratories will maintain a log of all unanticipated device-related 
complications leading to no test, such as absence of transport media, quantity not 
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6.0 
Statistics 
6.1 
6.2 
Study Design 
This is a cross-sectional, single visit study to evaluate the diagnostic accuracy of NAATs 
for the detection of NG and CT from pharyngeal and rectal swab specimens. Study 
participants will have swabs collected from both the rectum and pharynx, following any 
specimens taken for clinically indicated testing.  Each of the swabs from each anatomic 
location will be tested for both NG and CT using three different NAATs, with a 4th test 
collected and tested as a tiebreaker, if needed. [redacted] Diagnostic accuracy will be 
calculated with respect to an anatomic site infected status (ASIS) reference standard 
described in Section 3.5. As the reference standard is imperfect, we will report positive 
percent agreement and negative percent agreement in place of sensitivity and specificity. 
One potential limitation of this study is that all three of the tests in the reference standard 
are NAATs. However, all three tests [redacted]. So while, theoretically, there may be 
correlation in the errors produced by the three tests, the varied molecular targets and the 
different methods of capture, amplification and detection should mitigate this concern. 
Endpoints 
•
Infection status for Neisseria gonorrhoeae in the rectum as determined by each
NAAT
•
Infection status for Neisseria gonorrhoeae in the pharynx as determined by each
NAAT
•
Infection status for Chlamydia trachomatis in the rectum as determined by each
NAAT
•
Infection status for Chlamydia trachomatis in the pharynx as determined by each
NAAT
•
Anatomic Site Infection status as determined by the reference standard (as described
in Section 2.3)
6.3 
Sample Size 
For each assay, PPA and NPA35 will be estimated for each of three diagnostic tests 
defined by pathogen - anatomical site combinations: 
•
Neisseria gonorrhoeae in the rectum
•
Neisseria gonorrhoeae in the pharynx
•
Chlamydia trachomatis in the rectum
•
Chlamydia trachomatis in the pharynx
Each pathogen/anatomic site combination will be evaluated by constructing 95% Score 
confidence interval estimates for PPA and NPA and evaluating whether 90% can be ruled 
out by the lower bound of the CI with reasonable confidence.  

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NPA is expected to be greater than PPA and there will be considerably more not-infected 
than infected anatomic site, implying that there will be greater power for evaluating NPA. 
Thus it will suffice to size the study to evaluate PPA.  
Assuming that the true PPA is 96%,13 then 157, 174, and 196 evaluable participants that 
are infected are needed to have 80%, 85%, and 90% power respectively to demonstrate 
a PPA of greater than 90%. Since all of the participants will contribute to the evaluation of 
all three pathogen-anatomical site diagnostic tests, it will suffice to size the study for the 
diagnostic test with the lowest expected prevalence (except CT in the pharynx). The 
study will be more than sufficiently sized for more prevalent disease.  
It is expected that the disease prevalence of NG in the rectum, NG in the pharynx, and 
CT in the rectum will each be greater than 7.5% in the population under evaluation.13,14,16-
20 Table 3 summarizes the required number of participants to have 80% and 90% 
probabilities of obtaining the 157, 174, and 196 evaluable participants that are disease-
positive to have 80%, 85%, and 90% power when the true prevalence of disease is 7.5%, 
10%, and 12.5%.  
Table 3: Sample size required number to have 80% and 90% probabilities of obtaining the 157, 174, and 196 
evaluable participants that are disease-positive when the true prevalence of disease is 7.5%, 10%, and 12.5% in 
order to have 80%, 85%, and 90% power (two-sided alpha=0.05) to conclude that sensitivity is greater than 90% 
when the true sensitivity is 96%. 
6.4 
Randomization of Swabs 
Randomization will be employed to ensure that the swab order distribution is the same for 
4 tests, the three NAATs under investigation in the study and the tiebreaker test. SAS 
software (Cary, NC) will be used to generate permuted blocks with a block size of 24, to 
ensure that each NAAT and the tiebreaker test would have their swab distributions 
consisting of 1/4 being first swabs, 1/4 being second swabs, 1/4 being third swabs and 
1/4 being fourth swabs. 
6.5 
Monitoring 
The core protocol team will receive monthly reports monitoring study accrual, study 
conduct and completeness of data and swab collection. 
Prevalence of 
Disease 
Positive 
Power 
Required 
Disease Positive 
to Obtain Power 
Required Sample Size to 
have 80% probability of 
obtaining required 
Disease positive 
participants 
Required Sample Size to 
have 90% probability of 
obtaining required 
Disease positive 
participants 
7.5% 
80% 
157 
2250 
2325 
85% 
174 
2475 
2575 
90% 
196 
2775 
2875 
10% 
80% 
157 
1700 
1750 
85% 
174 
1860 
1925 
90% 
196 
2100 
2150 
12.5% 
80% 
157 
1350 
1400 
85% 
174 
1485 
1530 
90% 
196 
1665 
1715 

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The study will be reviewed by an independent statistician, who will not have an 
association with the protocol or device companies. The independent statistician will 
review the progress of the study. 
Additionally, the study statisticians will prepare reports for the independent statistician 
that will include baseline summaries (demographics and symptom status) and will 
address issues concerning the viability and appropriate execution of the protocol such as 
accrual, endpoint evaluability, and frequency of tests with equivocal results, invalid 
results, or no results. Issues with specimen collection will be summarized. The 
independent statistician will also review and evaluate the rates of disease. Toxicity is not 
expected given the nature of the study. 
The study cannot be stopped at the interim for reaching regulatory goals at this time to 
preserve error rates / coverage probability and ensure enough data for subgroup 
analyses. Since the trial cannot be stopped for attainment of the regulatory goal, no 
adjustment to confidence levels are necessary.  
Monitoring design assumptions for appropriate study sizing 
Infection rates will be evaluated to determine whether sample size adjustments are 
warranted. The sample size will not be adjusted based on the observed PPAs and NPAs, 
which will not be reviewed while the study is ongoing, but may be adjusted based only on 
the infection rates to ensure enough infected participants to estimate PPAs with desired 
precision. If infection is more prevalent than expected, a smaller sample size may be 
accepted. If however infection is rarer than anticipated, then increases to sample size will 
be considered. The infection rate will be evaluated approximately every 500 participants 
or every 3 months after the enrollment of the first participant, whichever comes first. 
6.6 
Analyses 
6.6.1 
Primary Analyses 
The result for each test will be compared with the ASIS for that anatomic site and 
organism.  For each diagnostic test, PPA and NPA will be estimated using 95% 
Score confidence intervals. Confidence intervals will be estimated using the 
Score method.34  
In the primary analysis, we will follow the FDA guidance document for 
incorporating indeterminate ASIS or test results for the test under consideration 
that are equivocal.34 Indeterminate ASIS results will be counted against the 
results for the test under consideration, as described in Tables 4 and 5. Tables 4 
and 5 will be presented in the analysis report, showing all combinations of 
Infected/Indeterminate/Not Infected with all outcomes from the test under 
consideration (Positive/Equivocal/Negative/No result or Invalid) 
If the test under consideration has “no result” because the test was not run and 
no attempt was made to test the sample, it will be excluded from the primary 
analysis. 
Table 4.  Use of the ASIS in calculation of the positive and negative percent agreement for 
the Company 2 Assay (Company 2) and Company 3 Assay (CT only). 
ASIS 
Infected 
Indeterminate* 
Not infected 

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Result of Test 
under 
Consideration** 
Positive 
A 
D 
G 
Equivocal 
B 
E 
H 
Negative 
C 
F 
I 
No result 
Exclude from analysis 
PPA 
A / (A+B+C+F) 
NPA 
I / (G + H + I + D) 
*The middle, indeterminate column will be included in the analyses as “Infected” or “Not infected” as
described in the sensitivity analyses below.
**Per the ASIS definition in Section 2.3
Comprehensive sensitivity analyses will be conducted considering multiple 
scenarios.  These include: 
•
Classify indeterminate tests on the basis of symptom status.  Include
indeterminate tests as infected if the participant is symptomatic in that
compartment.  Include indeterminate tests as not infected in the
participant is asymptomatic in that compartment.
•
Include all indeterminate tests as infected.
o
PPA=(A+D) / (A+B+C+D+E+F)
o
NPA=I / (G+H+I)
•
Include all indeterminate tests as not infected.
o
PPA=A / (A+B+C)
o
NPA=(F+I) / (D+E+F+G+H+I)
•
Considering indeterminate and equivocal test results as “missing”, with
the assumption of missing at random, and model the missing results.
•
Additional sensitivity analyses, including the use of latent class models,
will be explored.  Details will be in the statistical analysis plan.
Table 5. Use of the ASIS in calculation of the positive and negative percent agreement for 
the Company 1 Assay (Company 1)  and Company 3 Assay (NG only). 
ASIS 
Infected 
Indeterminate* Not infected 
Result of Test 
under 
Consideration** 
Positive/Detected 
A 
C 
E 
Negative/Not 
detected 
B 
D 
F 
No result 
Exclude from analysis 
PPA 
A / (A+B+D) 
NPA 
F / (C+E+F) 
*The middle, indeterminate column will be included in the analyses as “Infected” or “Not infected” as
described in the sensitivity analyses below.
**Per the ASIS definition in Section 2.3
If the test under consideration has “no result” because the test was not run and 
no attempt was made to test the sample or because of invalid ASIS results it will 
be excluded from the primary analysis. 
Comprehensive sensitivity analyses will be conducted.  These include: 
•
Classify indeterminate tests on the basis of symptom status.  Include
indeterminate tests as Infected if the participant is symptomatic in that
compartment.  Include indeterminate tests as not infected in the
participant is asymptomatic in that compartment.

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•
Include all indeterminate tests as infected
o
PPA = (A+C) / (A+B+C+D)
o
NPA = F / (E+F)
•
Include all indeterminate tests as not infected
o
PPA = A / (A+B)
o
NPA = (D+F) / (C+D+E+F)
•
Additional sensitivity analyses that consider indeterminate results as
“missing” and utilize imputation strategies, and graphical techniques
•
Additional sensitivity analyses, including the use of latent class models,
will be explored.  Details will be in the statistical analysis plan.
There are pros and cons to the manner in which indeterminates are handled and 
the resulting calculations of PPA and NPA. The primary definition is the most 
conservative but is biased downwards.  If PPA > 90% under this scenario, then 
the conclusion of PPA > 90% is clear. The second and third sensitivity analyses 
count indeterminates (treat all as infected or not infected, respectively) but may 
not be conservative for calculations of PPA and NPA.  
6.6.2 
Secondary Analyses 
Global Analyses 
For each diagnostic test: 
•
Positive and negative predictive values (PPV and NPV) will be
estimated. PPV and NPV will each be plotted as a function of prevalence
(point estimates and 95% pointwise confidence bands)
•
Positive and negative likelihood ratios will be estimated using 95% Score
confidence intervals
•
A sensitivity analysis will include calculating equivocal predictive values
and likelihood ratios
•
An additional sensitivity analysis will calculate PPV, NPV, and positive
and negative likelihood ratios for the sensitivity analyses where
indeterminates are counted as Infected and Not Infected
Subgroup Analyses 
For each of the three diagnostic tests, subgroup analyses will be conducted for 
males, females, symptomatic participants, and asymptomatic participants by 
pathogen and anatomic site. The 95% Score confidence interval estimates of 
PPA and NPA will be displayed using forest plots (one plot per test). The 95% 
Score confidence interval estimates of positive and negative likelihood ratios will 
be displayed using forest plots (one plot per test). PPVs and NPVs will each be 
plotted as a function of prevalence (point estimates and pointwise confidence 
bands). 
The sensitivity analyses where indeterminates are counted as Infected and Not 
Infected will be repeated for the four subgroup analyses. 
Exploratory Analysis 

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Developing methods for diagnostic benefit:risk analyses (e.g., diagnostic yield) will 
be applied. 
7.0 
Quality Control 
 
 
7.1 
Quality Controls and Study Monitoring 
 
The ARLG will provide direct access to the dataset for the purposes of monitoring and 
auditing by the DCRI, Harvard School of Public Health and inspection by local and 
regulatory authorities. The local site PI will ensure that study personnel are appropriately 
trained and applicable documentations are maintained.   
 
The DCRI will implement a Quality Plan to ensure that protocol training, data quality and 
data security are being undertaken.  
 
7.2 
Source Documents and Access to Source Data 
 
Source documents will not be removed from the sites.  Study monitoring will occur 
remotely, unless there are quality or performance concerns at a site, in which case an on-
site visit will occur. 
 
8.0 
Ethics / Protection of Human Subjects 
 
8.1 
Ethical Standards 
 
The investigator will ensure that the study will be conducted in accordance with all 
applicable national, regional, and local regulations.  
 
8.2 
Institutional Review Board and Informed Consent 
 
 
This protocol and any subsequent modifications must be reviewed and approved by the 
IRB responsible for oversight of the study, as listed on its FWA.  To minimize risks 
associated with loss of privacy and loss of confidentiality, oral informed consent will be 
obtained from the subject.  The informed consent process will describe the purpose of the 
study, the procedures to be followed, and the risks and benefits of participation.  A copy 
of a patient information form may be given to the subject (per local policy).   
 
8.3 
Subject Confidentiality 
 
Subjects will have code numbers and will not be identified by name. Subject 
confidentiality is strictly held in trust by the participating investigators, their staff, and the 
Sponsor(s) and their agents.   
 
The study protocol, documentation, data, and all other information generated will be held 
in strict confidence.  No information concerning the study or the data will be released to 
any unauthorized third party without prior written approval of the ARLG. 
 
All information provided by the ARLG and all data and information generated by the 
participating clinical site as part of the trial will be kept confidential by the site PI and 
other study personnel. This information and data will not be used by the site PI or other 
study personnel for any purpose other than conducting the trial. These restrictions do not 
apply to: (1) information which becomes publicly available through no fault of the site 
principal investigator or other study personnel; (2) information which is necessary to 

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disclose in confidence to an IRB solely for the evaluation of the trial (3) information which 
is necessary to disclose in order to provide appropriate medical care to a study subject; 
or (4) study results which may be published as described in Section 10.  
 
The study monitor or other authorized representatives of the sponsor may inspect all 
documents and records required to be maintained by the PI. The clinical study site will 
permit access to such records. 
 
 
All laboratory specimens, case report form data, reports and other records will be 
identified only by a coded number to maintain subject confidentiality.     
 
8.4 
Specimen Handling 
 
Specimens will be collected for this study and will be maintained by the ARLG at least 
until all companies have received their FDA approval for their assay.  
 
Use of samples for future research will be part of the consent process. There will not be 
any opt-in/opt-out language. Samples may be shared with other investigators at other 
institutions. No human genetic tests will be performed on samples. Each sample will be 
encoded (labeled) only with a barcode and a unique tracking number to protect subject’s 
confidentiality.  
 
There are no benefits to subjects in the collection, storage and subsequent research use 
of specimens. Because the subject enrollment will be done anonymously, reports about 
future research done with subject’s samples will NOT be reported into their health 
records.  
 
9.0 
Data Handling and Record Keeping 
 
 
9.1 
Data Collection 
 
This study will use eClinical OS (eCOS), a Web-based e-CRF database used by the 
DCRI.  The investigator's site staff who will be entering data will receive training on the 
system, after which each person will be issued a unique user identification ID and 
password. 
 
For security reasons, and in compliance with regulatory guidelines, it is imperative that 
only the persons who own the user IDs and passwords access the system using their 
own unique access codes. Access codes are nontransferable. Site personnel who have 
not undergone training may not use the system and will not be issued a user ID and 
password until appropriate training is completed. 
 
 
9.2 
Inspection of Records 
 
The DCRI will implement a Quality Plan to, at a minimum, ensure that activities 
proposed by the PI to ensure protocol training, data quality and data security are being 
undertaken. 
 
Source Documents and Access to Source Data 
 
Source documents will not be removed from the sites.  
 
 
9.3 
Retention of Records 

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Records will be maintained during the investigation and for a period of 2 years after the 
latter of the following dates: The date on which the investigation is terminated or 
completed, or the date that the records are no longer required for purposes of 
supporting a premarket approval application or a notice of completion of a product 
development protocol. Records may not be destroyed without the prior approval of the 
ARLG.  
 
 
9.4 
Confidentiality 
 
The study protocol, documentation, data, and all other information generated by this 
study will be maintained in a secure manner and will be kept confidential as required by 
law. 
 
The EDC database (eClinical OS data) used by the DCRI will be hosted by Merge which 
uses a Peak 10 datacenter located in Morrisville NC. Database access will be limited to 
study personnel who are issued a unique user identification and password. Data will 
be entered at each site by study personnel. No information concerning the study or the 
data will be released to any third party without prior written approval of the Sponsor. 
Study records may be reviewed in order to meet federal or state regulations. 
Reviewers may include the IRBs, the DCRI and the NIH. 
 
10.0 
Publication Policy  
 
Following completion of the study, the investigator may publish the results of this research in a 
scientific journal under the oversight of the Publication Committee of the ARLG.  
 
The ARLG Publication Committee comprises representatives of the network cores, thought-
leaders, statistics and data management center (SDMC), and is responsible for generation and 
coordination of the publications that report scientific findings of the network. All public 
presentations (abstracts, manuscripts, slides and text of oral or other presentations, and text of 
any transmission through any electronic media) by participating investigators, participating 
institutions, SDMC, and ARLG that use ARLG data and are intended to represent the ARLG or 
are supported by the ARLG will be reviewed by the Publication Committee per the Publication 
Committee charter and must include the following statement: “Research reported in this 
publication was supported by the National Institute Of Allergy And Infectious Diseases of the 
National Institutes of Health under Award Number UM1AI104681. The content is solely the 
responsibility of the authors and does not necessarily represent the official views of the National 
Institutes of Health.” 
 
The Publication Committee will ensure that the study results are presented by experts in the field 
that have working knowledge of the study design, implementation, data synthesis/analysis, and 
interpretation. The committee goals are to ensure that any confidential or proprietary information 
is protected, and that all appropriate statistical analyses have been included.  
 
The ARLG Publication Committee will adhere to the trials registration policy adopted by the 
International Committee of Medical Journal Editors (ICMJE) member journals. This policy 
requires that all applicable clinical trials be registered in a public trials registry such as 
ClinicalTrials.gov, which is sponsored by the National Library of Medicine. Other biomedical 
journals are considering adopting similar policies.  
 
In June 2007, the ICMJE adopted the WHO's definition of clinical trial: "any research study that 
prospectively assigns human participants or groups of humans to one or more health-related 
interventions to evaluate the effects on health outcomes."  

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•
Health-related interventions include any intervention used to modify a biomedical or
health-related outcome (for example, drugs, surgical procedures, devices, behavioral
treatments, dietary interventions, and process-of-care changes).
•
Health outcomes include any biomedical or health-related measures obtained in patients
or participants, including pharmacokinetic measures and adverse events.
All investigators funded by the NIH must submit or have submitted for them to the National 
Library of Medicine’s PubMed Central an electronic version of their final, peer-reviewed 
manuscripts upon acceptance for publication, to be made publicly available no later than 12 
months after the official date of publication. The NIH Public Access Policy ensures the public has 
access to the published results of NIH-funded research. It requires investigators to submit final 
peer-reviewed journal manuscripts that arise from NIH funds to the digital archive PubMed 
Central upon acceptance for publication. Further, the policy stipulates that these papers must be 
accessible to the public on PubMed Central no later than 12 months after publication.  
Refer to: http://publicaccess.nih.gov/ 

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11.0 
References 
1.
Company 1 Assay package insert. Vol. XXX, Rev .
2.
Company 2 Assay package insert. Vol. XXX Rev .
3.
Company 3 Assay package insert. Vol. Ref XXX .
4.
WHO. Global Incidence and prevalence of selected curable sexually transmitted 
infections.  (WHO Department of Reproductive Health and Research, 2008).
5.
CDC. Sexually Transmitted Disease Surveillance.  (2013).
6.
Liu, B., et al. Chlamydia and gonorrhoea infections and the risk of adverse obstetric 
outcomes: a retrospective cohort study. Sex Transm Infect 89, 672-678 (2013).
7.
O'Brien, J.P., Goldenberg, D.L. & Rice, P.A. Disseminated gonococcal infection: a 
prospective analysis of 49 patients and a review of pathophysiology and immune 
mechanisms. Medicine 62, 395-406 (1983).
8.
Westrom, L., Joesoef, R., Reynolds, G., Hagdu, A. & Thompson, S.E. Pelvic 
inflammatory disease and fertility. A cohort study of 1,844 women with laparoscopically 
verified disease and 657 control women with normal laparoscopic results. Sex Transm 
Dis 19, 185-192 (1992).
9.
Rottingen, J.A., Cameron, D.W. & Garnett, G.P. A systematic review of the 
epidemiologic interactions between classic sexually transmitted diseases and HIV: how 
much really is known? Sex Transm Dis 28, 579-597 (2001).
10.
Deguchi, T., Yasuda, M. & Ito, S. Management of pharyngeal gonorrhea is crucial to 
prevent the emergence and spread of antibiotic-resistant Neisseria gonorrhoeae. 
Antimicrob Agents Chemother. 56, 4039-4040 (2012).
11.
Workowski, K.A. & Bolan, G.A. Sexually transmitted diseases treatment guidelines, 
2015. MMWR Recomm Rep 64, 1-137 (2015).
12.
Zakher, B., Cantor, A.G., Pappas, M., Daeges, M. & Nelson, H.D. Screening for 
gonorrhea and chlamydia: a systematic review for the U.S. Preventive Services Task 
Force. Ann Intern Med 161, 884-893 (2014).
13.
Ota, K., Tamari, I. & Smieja, M. Detection of Neisseria gonorrhoeae and Chlamydia 
trachomatis in pharyngeal and rectal specimens using the BD ProbeTec ET system, the 
Gen-Probe Aptima Combo 2 assay and culture. Sexually Transmitted Infections 85, 
182-186 (2009).
14.
Schachter, J., Moncada, J. & Liska, S. Nucleic acid amplification tests in the diagnosis of 
chlamydial and gonococcal infections of the oropharyngx and rectum in men who have 
sex with men. Sexually Transmitted Diseases 35, 637-642 (2008).
15.
Trebach, J.D., Chaulk, C.P., Page, K.R., Tuddenham, S. & Ghanem, K.G. Neisseria 
gonorrhoeae and Chlamydia trachomatis among women reporting extragenital 
exposures. Sex Transm Dis 42, 233-239 (2015).
16.
Bachmann, L., Johnson, R. & Cheng, H. Nucleic acid amplications test for diagnosis of 
Neisseria gonorrhoeae and Chlamydia trachomatis rectal infections. Journal of Clinical 
Microbiology 48, 1827-1832 (2010).
17.
Cosentino, L.A., et al. Use of nucleic acid amplification testing for diagnosis of anorectal 
sexually transmitted infections. J Clin Microbiol 50, 2005-2008 (2012).
18.
Geelen, T.H., et al. Performance of cobas(R) 4800 and m2000 real-time assays for 
detection of Chlamydia trachomatis and Neisseria gonorrhoeae in rectal and self-
collected vaginal specimen. Diagn Microbiol Infect Dis 77, 101-105 (2013).
19.
Moncada, J., Schachter, J., Liska, S., Shayevich, C. & Klausner, J.D. Evaluation of self-
collected glans and rectal swabs from men who have sex with men for detection of

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Chlamydia trachomatis and Neisseria gonorrhoeae by use of nucleic acid amplification 
tests. J Clin Microbiol 47, 1657-1662 (2009). 
20.
Perry, M.D., Jones, R.N. & Corden, S.A. Is confirmatory testing of Roche cobas 4800
CT/NG test Neisseria gonorrhoeae positive samples required? Comparison of the Roche
cobas 4800 CT/NG test with an opa/pap duplex assay for the detection of N
gonorrhoeae. Sex Transm Infect 90, 303-308 (2014).
21.
Harryman, L., et al. Comparative performance of culture using swabs transported in
Amies medium and the Aptima Combo 2 nucleic acid amplification test in detection of
Neisseria gonorrhoeae from genital and extra-genital sites: a retrospective study. Sex
Transm Infect 88, 27-31 (2012).
22.
Walsh, A., Rourke, F.O. & Crowley, B. Molecular detection and confirmation of Neisseria
gonorrhoeae in urogenital and extragenital specimens using the Abbott CT/NG
RealTime assay and an in-house assay targeting the porA pseudogene. Eur J Clin
Microbiol Infect Dis 30, 561-567 (2011).
23.
Goldenberg, S., Finn, J., Sedudzi, E., White, J. & Tong, C. Performance of the
GenExpert CT/NG Assay compared to that of the Aptima AC2 Assay for Detection of
Rectal Chlamydia trachomatis and Neisseria gonorrhoeae by use of Residual Aptima
Samples. Journal of Clinical Microbiology 50, 3867-3869 (2012).
24.
Cosentino, L., et al. A Validation Study of the Cepheid Xpert CT/NG for detecting
Chlamydia trachomatis and Neisseria gonorrhoeae in Rectal Samples. Sex Transm
Infect 91, A1-A258 (2015).
25.
CDC. Recommendations for the Laboratory-Based Detection of Chlamydia trachomatis
and Neisseria gonorrhoeae.  (2014).
26.
Gen-Probe. APTIMA Chlamydia trachomatis Assay. Vol. 501799 Revision D (San Diego,
2012).
27.
Gen-Probe. APTIMA Neisseria gonorrhoeae Assay. Vol. 502486 Revision A 73 (San
Diego, 2011).
28.
Boyadzhyan, B., Yashina, T., Yatabe, J.H., Patnaik, M. & Hill, C.S. Comparison of the
APTIMA CT and GC assays with the APTIMA combo 2 assay, the Abbott LCx assay,
and direct fluorescent-antibody and culture assays for detection of Chlamydia
trachomatis and Neisseria gonorrhoeae. J Clin Microbiol 42, 3089-3093 (2004).
29.
Chernesky, M.A., et al. Ability of new APTIMA CT and APTIMA GC assays to detect
Chlamydia trachomatis and Neisseria gonorrhoeae in male urine and urethral swabs. J
Clin Microbiol 43, 127-131 (2005).
30.
Moncada, J., Donegan, E. & Schachter, J. Evaluation of CDC-recommended
approaches for confirmatory testing of positive Neisseria gonorrhoeae nucleic acid
amplification test results. J Clin Microbiol 46, 1614-1619 (2008).
31.
Tabrizi, S.N., et al. Evaluation of six commercial nucleic acid amplification tests for
detection of Neisseria gonorrhoeae and other Neisseria species. J Clin Microbiol 49,
3610-3615 (2011).
32.
BD. BD ProbeTec Chlamydia trachomatis (CT) Qx Amplified DNA Assay package insert.
Vol. 8081408 (2010).
33.
BD. BD ProbeTec Neisseria gonorrhoeae (GC) Qx Amplified DNA Assay package insert.
Vol. 8081409 (2010).
34.
FDA. Establishing the performance characteristics of in vivo diagnostics devices for
Chlamydia trachomatis and/or Neisseria gonorrhoeae: screening and diagnostic testing.
(2011).

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35. 
FDA. Statistical Guidance on Reporting Results from Studies Evaluating Diagnostic 
Tests. (2007). 
 
 

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APPENDIX A - Schedule of Events 
 
 
 
Enrollment 
Clinical Evaluations 
Review of eligibility 
X 
Verbal consent 
X 
Baseline CRFs 
X 
Collection of Specimens 
Rectal swabs 
X 
Pharyngeal swabs 
X