1. Chlamydia and Gonorrhea Lab Update þBurning Questions þLaboratory Guidelines þCT Immunobiology Consultation The findings and conclusions in this presentation are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention

2. Alternate Specimen Verification þRectal and oropharyngeal swabs  FDA clearance being pursued  Establish an external specimen bank þHome collected vaginal swabs  Interest but studies have not commenced (that I’m aware of)

3. þPerformance þDiscreet packaging  Must meet federal regulations for shipping diagnostic specimens  Wet vs dry swabs þGreater temperature ranges than covered in the product insert þNOT FDA CLEARED  What are the parameters for an off-label verification study?  Need sound guidance Considerations Self-Obtained Vaginal Swabs in a Home Setting

4. Sensitivity of Different Specimens by Three Different Assays AssayFCUCxS-vagC-vag TMA72%89%93%90% PCR84%91% 93% LCR98%96%98%100% Combined81%91%93% FCU – first catch urine, Cx – endocervix, S-vag – self-collected vaginal swab, C-vag - Clinician collected vaginal swab. Schachter J, et al. JCM 41;2003:3784

5. Preliminary Vaginal Swab Mailing Kit

6. One Example of a Vaginal Swab Mailing Kit (Gen-Probe ® Vaginal Swab Transport Tube, Vaginal Swab, Inner Medical Specimen Transport Tube, Vaginal Swab Collection Instructions, Follow-up Questionnaire, and Mailing Can) Mailing Can Follow-up Questionnaire Swab

7. Swab Specimen Transport Conditions for Commercially Available CT/GC NAATs TestSwab ConditionTransport Temperature Time to Test BD ProbeTecDry (no liquid media)2 to 27 o C4 to 6 days GenProbe APTIMA (vaginal swabs FDA cleared) Penetrable top tube containing specimen buffer 2 to 30 o CUp to 60 days Roche AmplicorTubes containing Chlamydia culture medium (eg. M4, 2ST, Bartels, etc) 18 to 25 o CWithin 1 hour 2 to 8 o CUp to 7 days Less than -20 o CUp to 30 days

8. Package Temperature during USPS Transit from St. Louis to New Orleans Package placed in St. Louis USPS drop box July 22, 2005 and received at 2:30pm July 26, 2005 in New Orleans. Temperatures were recorded at 10 minute intervals.

9. Package Temperature during USPS Transit from Jackson to New Orleans Package placed in Jackson USPS drop box July 29, 2005 and received at 2:00pm August 10, 2005 in New Orleans. Temperatures were recorded at 10 minute intervals.

10. Temperature recorded at 1 minute intervals August 21, 2005.

11. APTIMA Test Result for Chlamydia trachomatis (Relative Light Units) Initial Temperature / Duration Final Temperature / Duration Inoculum (Inclusion forming units) 24 o C / 24 hours37 o C / 24 hours 1011041102 10 3 12261115 10 5 11951087 In Vitro Assessment of Specimen Stability

12. APTIMA Test Result for Chlamydia trachomatis (Relative Light Units) Initial Temperature / Duration Final Temperature / Duration Inoculum (Inclusion forming units) 37 o C / 24 hours56 o C / 24 hours 1011721124 10 3 11331114 10 5 11421115 In Vitro Assessment of Specimen Stability

13. APTIMA Test Result for Chlamydia trachomatis (Relative Light Units) Initial Temperature / Duration Final Temperature / Duration Inoculum (Inclusion forming units) 56 o C / 24 hours24 o C / 24 hours 1010591023 10 3 12111951 10 5 11471113 In Vitro Assessment of Specimen Stability

14. Alternate Specimen Verification þRectal and oropharyngeal swabs  FDA clearance being pursued  Establish an external specimen bank þHome collected vaginal swabs  Interest but studies have not commenced (that I’m aware of) þAPHL / CDC STD Steering Committee Workgroups to develop Verification Protocols  Involve CMS (CLIA)  Part of revised CDC Laboratory guidelines

16. LGV Testing þSpecimens submitted to CDC in 2007 for the detection of LGV  ompA genotyping (not serology) Specimen TypeNumber Submitted (%) Results Unable to amplify (%) CT Negative (%)CT Positive (%)LGV (%) Rectal Swab52 (85%)8 (15%)14 (27%)25 (48%)5 (10% of all or 20% of CT+) Urethral Swab7 (11%)3 (43%)04 (57%)0 Urine1 (2%)0010 Unknown1 (2%)0010

17. þImportance of differentiating LGV from non-LGV rectal CT infections  Course of recommended Tx varies  No commercial differential test available  Seems that settings that have been testing for rectal CT infections and managing those infections as non-LGV have not seen increases in proctocolitis Refer to Dr. Mark Pandori’s presentation at the 2008 STD Prevention meeting  Sx and Asx infections reported  CDC does not have a survelliance program LGV Submission of specimens for reference testing should be based on local descions þSerology for rectal LGV testing  No data LGV Testing

18. Laboratory Guidelines

19. Background ü1982  Laboratory Diagnosis of Chlamydia trachomatis Infections  Isolation and serology ü1993  Recommendations for the Prevention and Management of Chlamydia trachomatis Infections Isolation Non-culture tests  Immunoassays, nucleic acid hybridization tests Screening Sexual assault Presumptive Laboratory Diagnosis

20. Screening Tests to Detect Chlamydia trachomatis and Neisseria gonorrhoeae Infections - 2002 üGuideline Development  CDC identified questions  reviewed published literature from 1990 - 2000  prepared tables of evidence  drafted recommendations  External consultants asked to review the drafted recommendations selected on the basis of expertise and/or disciplinary or organizational affiliations  CDC considered all suggestions from external consultants and finalized recommendations

21. Revising Laboratory Guidelines for the Detection of Chlamydia trachomatis and Neisseria gonorrhoeae Infections üSimilar format of previous guidelines / consultations  Convene an expert review panel  Identify / refine key questions  Review literature  External consultants asked to review the drafted recommendations selected on the basis of expertise and/or disciplinary or organizational affiliations  CDC considered all suggestions from external consultants and finalized recommendations

22. Issues þWhat is the sensitivity and specificity of available tests for CT and GC  Is it correct to lump NAATs þDoes the sensitivity and specificity of available for CT and GC vary with respect to the anatomic site from which the specimen was collected and/or the specimen type þWhat class of tests should be recommended for the detection of CT and GC infection; stratify by anatomic site and/or specimen type þWhat test or combination of tests should be recommended confirmation of CT and GC infection

23. Issues þCan routine supplemental testing improve the PPV of certain CT and GC tests and at what prevalence level should it be recommended

24. Positive Predictive Value likelihood that the test accurately predicts the true infection status of the person tested Example Test 1,000 persons Test Specificity99.6 % True Positive 100False Positive 4 Prevalence of Infection  HIV, syphilis, chlamydia, gonorrhea, etc 10 % Positive Predictive Value 100/104 = 96 % True Positive 4False Positive 4 Prevalence of Infection  HIV, syphilis, chlamydia, gonorrhea, etc 0.4 % Positive Predictive Value 4/8 = 50 %

25. Example Test 1,000 persons Test Specificity99.6 % True Positive 100False Positive 4 Prevalence of Infection  HIV, syphilis, chlamydia, gonorrhea, etc 10 % Positive Predictive Value 100/104 = 96 % True Positive 4False Positive 4 Prevalence of Infection  HIV, syphilis, chlamydia, gonorrhea, etc 0.4 % Positive Predictive Value 4/8 = 50 % Positive Predictive Value likelihood that the test accurately predicts the true infection status of the person tested

26. Issues þCan routine supplemental testing improve the PPV of certain CT and GC tests and at what prevalence level should it be recommended þDoes pooling urine specimens diminish NAAT performance þShould recommendations be made on testing in situations involving medicolegal issues including adult and pediatric CT and GC infections (note: making a recommendation of any test beyond culture would require a preliminary discussion recommendations for off-label testing)

27. Issues þWhat test(s) should be recommended for the laboratory diagnosis of LGV and would these vary based on the presentation (i.e. inguinal versus anorectal presentation) þWhat serologic cut-off values should be used if serology is recommended to aid LGV diagnosis þWhat (if any) recommendations be made concerning in vitro antibiotic susceptibility testing for GC including but not limited to the actual methodology for the procedure

28. Chlamydia Immunology and Control Expert Advisory Meeting Atlanta, April 23-25, 2008 þHighlight the key questions related to chlamydia natural history, pathogenesis, and immunobiology that have the most important implications for control of chlamydia and its sequelae þAssess extent to which existing data address these key questions, especially with respect to their potential relevance to prevention programs þIdentify important remaining gaps in knowledge that would have implications for prevention

29. Chlamydia trachomatis þPathology is primarily a function of the host-response  Little intrinsic toxicity þProtective immunity has been demonstrated in animal models or animals naturally infected with related chlamydiae  Mice, guinea pigs, sheep etc  Th1 type DTH in mice þSusceptibility to reinfection is common  High rates of reinfection  Early Tx of mice resulted in a disruption of immunity  Th2 type DTH in mice þ Development of pathology poorly understood  How long?  Predisposing factors?  Reinfection or persistent infection? þ Development of immunity that protects against subsequent CT infections has not been conclusively documented in humans  Commercial sex workers seem to be refractory to infection after time þ Human data is lacking (absent) on the mechanisms associated with reinfection  Bacterial?  Host? Some of what we knowSome of what we don’t know

30. Chlamydia trachomatis þScreening and Tx reduce the natural duration of infection þDespite screening, reinfection rates seem to be increasing  Brunham et al JID 2005 þAntibiotic resistance has not been demonstrated among human CT isolates  It has been shown in pig isolates þ How long does CT persist if left untreated?  Brazilian data suggests most half of the infections are cleared after a year and all are cleared by 5 years þ What are the reasons for increases in reinfection rates?  Better Dx tests  Increased / more targeted screening  Bacterial changes þ Why are some CT infections more difficult to treat?  Golden et al NEJM 2005 Some of what we knowSome of what we don’t know

31. Some of What we Learned þGood evidence that repeated episodes of acute diagnosed PID increase the risk of tubal infertility þCT related damage is primarily a function of the host response þUnderstanding of pathogenesis or timing of occurrence of damage is more limited than most realized  additional work needed if new approaches to prevention are to be developed  Existing animal model data offer somewhat less insight than expected but alternative approaches possible þData addressing the mechanisms, development, or role of immunity in humans are quite limited  But approaches for addressing these gaps provided