Susanne Norris Zanto, MPH, MLS (ASCP) CM, SM Montana Public Health Laboratory
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1 Susanne Norris Zanto, MPH, MLS (ASCP) CM, SM Montana Public Health Laboratory
2 Describe the challenges in syphilis diagnostics Present two testing algorithms Non-treponemal test as initial screen Treponemal test as initial screen Discuss how guidelines will impact practices in public health and clinical laboratories
3 Syphilis Infection 6 weeks to 6 months ~ 18 months Many years to a lifetime Infection Primary Chancre Secondary Rash Latent Asymptomatic Tertiary Benign gumatous lesions Cardio-vascular syphilis Neurosyphilis Incubation period 9 90 days Early Syphilis 1 2 syears Late Syphilis Many years to a lifetime
4 In 2000, rate was lowest since 1941 Re-emerged as Public Health threat in 2001 CDC - STD Surveillance, 2009
5 Majority of cases reported in MSM In 2009, 62% of P&S cases 4% in 2000 P & S Reported Cases in 2009, CDC - STD Surveillance, 2009
6 Rates among women increased 36% from , but decreased 6.7% during P&S syphilis increased 4% among men during CDC - STD Surveillance, 2009
7 Efforts to eliminate syphilis are falling short Current state of laboratory diagnostics is contributing factor Limited access to direct detection Heavy reliance on serologically based tests Indirect evidence Requires normal immune response (HIV) Non-specific and false negative reactions
8 CDC guidelines influence both public health laboratory and clinical laboratory practices Address appropriate use of tests Test Performance Dependent on stage of Syphilis Infection? What about HIV co-infection? Test Interpretation / Need for Reflex testing Need for repeat testing, additional specimens
9 Expert consultation held in January 2009 Team met by conference call to continue consensus process Will most likely be divided into four sections Direct Detection of T. pallidum(tp) Congenital Syphilis Neurosyphilis Serology Anticipate publication late 2011, early 2012
10 Darkfieldmicroscopy Role in high prevalence sites Serology still recommended Immunostaining High sensitivity and specificity in ID of primary syphilis lesions Nucleic Acid Amplification tests Useful test in lesions of Primary Syphilis Usefulness in secondary syphilis unclear Is PCR sensitive enough to identify Tp in blood, serum, plasma or CSF? Quality Assurance and PT needed
11 IgMshould be used in concert with Tpdirect detection (immunostaining, PCR, DF) A reactive IgMtest is useful Non-reactive IgMtest does not rule out congenital syphilis A four-fold or greater ratio of neonatal to maternal titers is rarely useful Treatment decisions should not be based on laboratory results
12 Neurosyphilis cannot be diagnosed serologically Serum RPR > 1:32 can predict which asymptomatic individuals are most likely to have CSF findings consistent with neurosyphilis The use of VDRL in evaluating CSF may still be worthwhile WBC cutoffs, TP-PA of 1:320, CXCL13 show promise Protein levels and TPHA titer index are not useful tests for evaluating CSF
13 Considerations Prevalence of disease in population Performance characteristics of test Purpose of the test (screening, confirmation, disease management) Subjectivity of the test/experience of the technologist Reflex testing needs Need for capital equipment Automation needs Technical requirements Cost
14 Most EIAs perform equally well for screening Those detecting both IgMand IgGappear more sensitive in early disease Limited head-to-head comparison data especially newer FDA approved assays There is variability in the performance of different treponemal tests Due to antigens, conjugates and methods The overall agreement between treponemaltests is usually >95%, even in 1 disease TP-PA appears to be the more sensitive than FTA-ABS
15 Non-treponemal tests RPR, VDRL, TRUST Treponemal tests FTA-ABS, TP-PA (Confirmatory Assays) EIA, CLIA, MIBA (Screening Assays) Rapid (Point of Care) tests Recent FDA approval of a treponemaltest
16 RPR Rapid Plasma Reagin VDRL Venereal Disease Research Laboratory TRUST ToluidineRed Unheated Serum Test
17 Antigen suspension is a carbon particle cardiolipinlecithin-coated cholesterol antigen Detects reagin, an antibody-like substance present in serum or plasma The reaginbinds to the test antigen, causing macroscopic flocculation Qualitative and Quantitative
18 Another test that detects reagin Qualitative and Quantitative slideflocculation test Requires heat inactivated serum Approved for CSF testing
19 Identical to RPR, but uses a dye instead of charcoal for visualization
20 PROS High Sensitivity Low cost Does not detect past infections Requires little equipment for testing Usually requires only one reflex test Useful for treatment monitoring CONS Lower specificity Labor-intensive Subjective results Manual data manipulations
21 EIA Enzyme Immunoassay CLIA Chemiluminescence Immunoassay MIBA Micro Immunobead Assay FTA-ABS Fluorescent Treponemal Antibody Absorbed TP-PA Treponema pallidum Particle Agglutination
22 Trep-Check Antihuman IgGantibodies conjugated with horseradish peroxidase(hrpo). Trep-Sure Horseradish Peroxidase(HRPO) conjugated treponemalantigens Lower percentage of false-positive results compared to Trep-Check Captia-Syphilis
23 Total Antibody to T. pallidum(iggand IgM) Fully automated chemiluminescence immunoassay Architect Liaison
24 Fully-automated, random access multiplex assay Multiplex Bead Technology Detects IgGto T. pallidum15kda, 17kDa, 47kDa (recombinant proteins) Also has separate IgM assay BioPlex 2200
25 PROS High Sensitivity High Specificity Objective results Automation / high throughput Interface with LIS CONS Cannot distinguish between active and previously treated disease Potential for over diagnosis and over treatment May require more resources for EPI/DIS investigations Specific, potentially costly instrumentation May require multiple reflex tests for resolving discrepants
26 Semi-quantitative graded 1+ to 4+ Even with absorbant, lower specificity than other treponemaltests and probably lower sensitivity
27 Qualitative gelatinparticleagglutination assay Reportedas Reactive, Non-Reactive, Inconclusive or Indeterminate
28 The selection of specific tests for screening depends on the setting, the population and the individual patient More than one algorithm will be necessary One using a non-treponemal test as initial screen One using a treponemaltest as initial screen A combination of treponemal and non-treponemal tests must be used; a single treponemaltest cannot be relied upon for syphilis diagnosis
29 Traditional Testing Algorithm Using Non-Treponemal Initial Screen Non-Treponemal Test RPR, VDRL, TRUST Reactive Titer Nonreactive Not syphilis (or early syphilis) Treponemal Test FTA-ABS, TP-PA Reactive Syphilis - Treat Nonreactive False positive Non-Treponemal Test Pope Infect Med 2004
30 Testing Algorithm Using EIA, CIA, MIBA as Initial Screen (Reverse Sequence) A1+ A2 Quantitative Nontreponemal (i.e. RPR) A1+ A2+ Consistent with Syphilis (past or current infection) A1 Syphilis EIA, CIA, or MIBA A1- Negative for Syphilis antibodies A1+ A2- A3 Treponemal Test that uses a different antigen or platform from A1 (i.e. TP-PA) * Laboratory should report the results of all three assays (if applicable) within 7 days A1+ A2- A3+ Possible Syphilis infection; Requires further historical and clinical evaluation A1+ A2- A3- Unconfirmed EIA; Unlikely to be Syphilis; If patient is at risk for syphilis, re- test in 1 month
31 There is no apparent advantage to performing quantitativetreponemaltests There is insufficient data to definitively recommend IgMtests for at this time (noncongenital) May have a role in resolving a reactive treponemal screening test with a non-reactive non-treponemal test
32 Serologic response following successful treatment of syphilis infection is unclear Most biological false positive reactions (both non-treponemaland treponemal) are seen in the sera of healthy individuals Data generally does not support that pregnant women have a higher rate of BFPs High rates in elderly may point to periodontal disease as a cause of BFPs
33 May be a role in areas where prevalence is high and where immediate treatment is the overriding concern due to a likely lack of follow up care May be a role as the additional treponemal test in the reverse sequence algorithm Usefulness in low prevalence populations unknown
34 Syphilis Health Check TM Recently FDA approved Moderately complex Seeking Waived Status Seeking Waived Status Treponemaltest only
35 Simultaneous yet separate detection of both treponemal and non-treponemal markers Not available in the U.S. (yet)
36 Comprehensive studies to compare treponemal and non-treponemal assays (currently FDAapproved) using sera from all stages of syphilis Need a serum bank of well characterized sera differentiated by clinical stage including coinfected HIV sera Studies to determine cause of BFPs
37 Studies to determine how soon after exposure a person can be screened, or time frame for rescreening if initial test is negative Need additional data about the persistence of IgM and the usefulness of IgM serology tests Usefulness of rapid point of care syphilis tests
38 No testing algorithm can take the place of clinical judgment Test results must be used in conjunction with the patient s clinical symptoms, medical and sexual history, and other clinical and/or laboratory findings to produce an overall clinical diagnosis.
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