Chlamydia and Gonorrhea Lab Update Burning Questions Laboratory

Chlamydia and Gonorrhea Lab Update þ Burning Questions þ Laboratory Guidelines þ CT 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

Alternate Specimen Verification þ Rectal and oropharyngeal swabs FDA clearance being pursued Establish an external specimen bank þ Home collected vaginal swabs Interest but studies have not commenced (that I’m aware of)

Self-Obtained Vaginal Swabs in a Home Setting Considerations þ Performance þ Discreet packaging Must meet federal regulations for shipping diagnostic specimens Wet vs dry swabs þ Greater temperature ranges than covered in the product insert þ NOT FDA CLEARED What are the parameters for an off-label verification study? Need sound guidance

Sensitivity of Different Specimens by Three Different Assays Assay FCU Cx S-vag C-vag TMA 72% 89% 93% 90% PCR 84% 91% 93% LCR 98% 96% 98% 100% Combined 81% 93% 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

Preliminary Vaginal Swab Mailing Kit

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

Swab Specimen Transport Conditions for Commercially Available CT/GC NAATs Test Swab Condition Transport Temperature Time to Test BD Probe. Tec Dry (no liquid media) 2 to 27 o. C 4 to 6 days Gen. Probe APTIMA Penetrable top tube (vaginal swabs FDA containing specimen cleared) buffer 2 to 30 o. C Up to 60 days Roche Amplicor 18 to 25 o. C Within 1 hour 2 to 8 o. C Up to 7 days Less than -20 o. C Up to 30 days Tubes containing Chlamydia culture medium (eg. M 4, 2 ST, Bartels, etc)

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: 30 pm July 26, 2005 in New Orleans. Temperatures were recorded at 10 minute intervals.

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

Temperature recorded at 1 minute intervals August 21, 2005.

In Vitro Assessment of Specimen Stability APTIMA Test Result for Chlamydia trachomatis (Relative Light Units) Initial Temperature / Duration Final Temperature / Duration Inoculum (Inclusion forming units) 24 o. C / 24 hours 37 o. C / 24 hours 10 1104 1102 103 1226 1115 105 1195 1087

In Vitro Assessment of Specimen Stability APTIMA Test Result for Chlamydia trachomatis (Relative Light Units) Initial Temperature / Duration Final Temperature / Duration Inoculum (Inclusion forming units) 37 o. C / 24 hours 56 o. C / 24 hours 10 1172 1124 103 1133 1114 105 1142 1115

In Vitro Assessment of Specimen Stability APTIMA Test Result for Chlamydia trachomatis (Relative Light Units) Initial Temperature / Duration Final Temperature / Duration Inoculum (Inclusion forming units) 56 o. C / 24 hours 24 o. C / 24 hours 10 1059 1023 103 1211 1951 105 1147 1113

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

LGV Testing þ Specimens submitted to CDC in 2007 for the detection of LGV omp. A genotyping (not serology) Specimen Type Number Submitted (%) Results Unable to amplify (%) CT Negative (%) CT Positive (%) LGV (%) Rectal Swab 52 (85%) 8 (15%) 14 (27%) 25 (48%) 5 (10% of all or 20% of CT+) Urethral Swab 7 (11%) 3 (43%) 0 4 (57%) 0 Urine 1 (2%) 0 0 1 0 Unknown 1 (2%) 0 0 1 0

LGV Testing þ Importance 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 þ Serology for rectal LGV testing No data

Laboratory Guidelines

Background ü 1982 Laboratory Diagnosis of Chlamydia trachomatis Infections Ø Isolation and serology ü 1993 Recommendations for the Prevention and Management of Chlamydia trachomatis Infections • Isolation • Non-culture tests Ø Immunoassays, nucleic acid hybridization tests • Screening • Sexual assault • Presumptive Laboratory Diagnosis

Screening Tests to Detect Chlamydia trachomatis and Neisseria gonorrhoeae Infections - 2002 ü Guideline 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

Revising Laboratory Guidelines for the Detection of Chlamydia trachomatis and Neisseria gonorrhoeae Infections ü Similar 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

Issues þ What is the sensitivity and specificity of available tests for CT and GC Is it correct to lump NAATs þ Does 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 þ What class of tests should be recommended for the detection of CT and GC infection; stratify by anatomic site and/or specimen type þ What test or combination of tests should be recommended confirmation of CT and GC infection

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

Positive Predictive Value likelihood that the test accurately predicts the true infection status of the person tested Example Test 1, 000 persons Test Specificity 99. 6 % Prevalence of Infection 10 % 0. 4 % HIV, syphilis, chlamydia, gonorrhea, etc True Positive 100 4 Positive Predictive Value False Positive 100/104 4/8 = 50 = %96 % 4

Positive Predictive Value likelihood that the test accurately predicts the true infection status of the person tested Example Test 1, 000 persons Test Specificity 99. 6 % Prevalence of Infection 10 % HIV, syphilis, chlamydia, gonorrhea, etc True Positive 100 Positive Predictive Value Prevalence of Infection False Positive 4 100/104 = 96 % 0. 4 % HIV, syphilis, chlamydia, gonorrhea, etc True Positive 4 Positive Predictive Value False Positive 4/8 = 50 % 4

Issues þ Can routine supplemental testing improve the PPV of certain CT and GC tests and at what prevalence level should it be recommended þ Does pooling urine specimens diminish NAAT performance þ Should 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)

Issues þ What 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) þ What serologic cut-off values should be used if serology is recommended to aid LGV diagnosis þ What (if any) recommendations be made concerning in vitro antibiotic susceptibility testing for GC including but not limited to the actual methodology for the procedure

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

Chlamydia trachomatis Some of what we know þ Pathology is primarily a function of the host-response Little intrinsic toxicity þ Protective immunity has been demonstrated in animal models or animals naturally infected with related chlamydiae Mice, guinea pigs, sheep etc Th 1 type DTH in mice þ Susceptibility to reinfection is common High rates of reinfection Early Tx of mice resulted in a disruption of immunity Th 2 type DTH in mice Some of what we don’t know 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?

Chlamydia trachomatis Some of what we know þ Screening and Tx reduce the natural duration of infection Some of what we don’t know 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 Despite screening, reinfection rates seem to be increasing Brunham et al JID 2005 þ What are the reasons for increases in reinfection rates? Better Dx tests Increased / more targeted screening Bacterial changes þ Antibiotic resistance has not been demonstrated among human CT isolates It has been shown in pig isolates þ Why are some CT infections more difficult to treat? Golden et al NEJM 2005

Some of What we Learned þ Good evidence that repeated episodes of acute diagnosed PID increase the risk of tubal infertility þ CT related damage is primarily a function of the host response þ Understanding 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 þ Data addressing the mechanisms, development, or role of immunity in humans are quite limited But approaches for addressing these gaps provided