New Approaches for Infectious New Approaches for Infectious Diseases Testing in Clinical Diseases Testing in Clinical LaboratoriesLaboratories

William M. Janda, Ph.D., D(ABMM)Professor Emeritus of PathologyUniversity of Illinois College of MedicineUniversity of Illinois at ChicagoDirector, Clinical Microbiology LaboratoryJohn H. Stroger, Jr. Hospital/Cook County Hospitals and Healthcare SystemChicago, Illinois

Topics to be AddressedTopics to be Addressed• New CDC HIV-1/2 testing algorithm

• Reverse-sequence testing for syphilis

• Use of nucleic acid amplification tests (NAATs) for diagnosis of extra-genital gonococcal/chlamydial infections

• Use of NAATS for documentation of child sexual abuse

Typical Course of HIV InfectionTypical Course of HIV Infection

HIV Markers Early in HIV Infection: Window HIV Markers Early in HIV Infection: Window Periods of Immunoassays (IA’s): First Through Periods of Immunoassays (IA’s): First Through Fourth GenerationFourth Generation

HIV-1/2 Testing: The HIV-1/2 Testing: The Former AlgorithmFormer Algorithm• Performance of a HIV-1/2 antibody

immunoassay (IA) on a patient serum specimen

• Reactive serum specimens are retested in duplicate using the same IA

• Repeatedly reactive serum specimens are tested by a confirmatory test• HIV-1 Western immunoblot →• Indirect fluorescent antibody test

• Cannot detect acute HIV-1 infection

CDC Criteria for CDC Criteria for HIV-1 Western HIV-1 Western Immunoblot Immunoblot InterpretationInterpretation

• Positive blot criteria • 1 →→→ Ab to

gp160/120 and p24

• 2 →→→ Ab to gp41 and p24

• 3 →→→ Ab to

gp160/120 and gp41

• No bands → Negative

• Any other combination→ Indeterminant

HIV Antibody ImmunoassaysHIV Antibody ImmunoassaysGeneration of HIV-1 assay

First generation •Used either viral lysate antigens•Detected only IgG-class antibodies

Second generation •Used either recombinant or synthetic peptides as antigens•Detected only IgG-class antibodies

Third generation Use synthetic peptides or recombinant antigensDetect both IgG- and IgM-class antibodies

Fourth generation Use synthetic peptides or recombinant antigensDetect IgG- and IgM-class antibodies Detect p24 (core) antigen

Third-Generation EIAsThird-Generation EIAs• “Sandwich” technique with enzyme-coupled HIV antigens

• Take advantage of the bi-/multivalent nature of antibodies → Improved specificity

• Only antibodies bound to ELISA wells that also bind HIV antigens generate a signal

• Nonspecifically bound antibodies less likely to bind HIV antigens

• Technology expands subtypes of detected antibodies• In direct ELISAs the conjugate is directed against a specific

antibody subtype (e.g., IgG), whereas sandwich technology permits detection of any antibody class, including IgM• Sandwich ELISA thus increases the ability to detect HIV

antibodies early in HIV infection (e.g., IgM, IgA)

Comparison of First (A) and Comparison of First (A) and Third-Generation (B) HIV EIA’sThird-Generation (B) HIV EIA’s

p24 Antigen p24 Antigen DetectionDetection

• Assay to detect the presence of the HIV viral core protein p24 in serum and plasma introduced in 1989

• Proved useful during early infection and improved detection of recent infection

• Used with antigen-antibody dissociation techniques as an aid to diagnosis of HIV infection in infants (Women Infants Transmission Study [WITS])

• p24 antigen detection kits approved by the FDA in 1989

Fourth Generation HIV-1/2 EIA Fourth Generation HIV-1/2 EIA (courtesy, BioRad)(courtesy, BioRad)

•New New HIV-1/2 HIV-1/2 Testing Testing AlgorithmAlgorithm

Includes 3 categories of tests:

4th generation HIV-1/2 Ab and Ag combo assay

HIV-1/HIV-2 discrimination assay

Nucleic acid testing

Multispot HIV-Multispot HIV-1/HIV-2 Rapid 1/HIV-2 Rapid Test (Bio-Rad)Test (Bio-Rad)

• Approved by the FDA in March, 2013 as a supplemental test

• Recommended option by CLSI

• Fast TAT of third- and fourth-generation immunoassays (<1 hour) and the Multispot test enables definitive same-day test results

Multi-Spot HIV-1/2 Rapid TestMulti-Spot HIV-1/2 Rapid Test• 1. Procedural control (goat anti-human IgG)• 2. HIV-2 peptide

• Peptide representing the immunodominant epitope of HIV-2 (gp36 envelope glycoprotein

• 3. Recombinant HIV-1 envelope glycoprotein• Recombinant gp41 expressed in E. coli (gp41 rDNA)

• 4. HIV-1 peptide• Peptide representing the immunodominant epitope of HIV-1 gp41

envelope glycoprotein

A1: GS HIV Combo Ag/Ab EIA

A2

Multispot *Multispot *

A1(-)

Negative for HIV-1 and HIV-2 antibodies and p24

Ag

A1+

HIV-1 +

HIV-1 antibodies

detected

Initiate care

(and viral load)

HIV-2 + HIV-2

antibodies detected

Initiate care

HIV-1&2 (-)

NAAT

NAAT+ Acute HIV-1 infection

Initiate care

NAAT (-) Negative for HIV-1

New CDC HIV-1/2 Testing AlgorithmNew CDC HIV-1/2 Testing Algorithm

*Multispot does not have confirmatory claim but must be used as a differentiating test

Multispot - InterpretationMultispot - Interpretation

• Nonreactive →→

• HIV-2 Reactive→

• HIV-1 Reactive →

• HIV Reactive →→• Undifferentiated

• INVALID →→→→

Confirmatory Confirmatory Tests:Transcription-Tests:Transcription-Mediated Mediated AmplificationAmplification

• RNA converted to DNA by reverse transcription

• DNA used as a transcription template (DNA to RNA)

• RNA reverse transcribed back to DNA

• RNA detected by probes• 30 copy/ml sensitivity• APTIMA HIV-1

qualitative assay (Gen-Probe) cleared for diagnostic use

Arizona DOHS/Maricopa Integrated Arizona DOHS/Maricopa Integrated Health Systems StudyHealth Systems StudyMMWR 2013;62:489-494MMWR 2013;62:489-494

• Screened all adult ED patients ages 18-64 years, July 2011-Feb 2013• Fourth-generation EIA used for screening with reflex to

Multispot (MS and WB)• Specimens negative by MS or WB tested for HIV-1 RNA

• Results• Detected previously undiagnosed HIV infection in 37

patients• 25 diagnoses were positive by MS, WB, or both • For 12 of the 37 patients, infection established by negative

MS and/or WB results and positive for HIV-1 RNA • Median HIV-1 viral loads in patients with acute infection was

3,636,176 copies/ml

Benefits to Identifying Acute Benefits to Identifying Acute HIV-1 InfectionHIV-1 Infection• Acute infection accounts for 5-10% of HIV infection among

those tested• Risk of transmission from persons with early infection is higher

than from those with established infections• Persons who have been infected for less than 6 months

account for almost 50% of all onwards transmission of HIV• Enables intervention to interrupt transmission

• Persons with acute HIV infection named 2.5 times as many partners

• Persons with acute HIV infection had nearly twice as many partners with undiagnosed HIV infection as did persons with established infections

Implications for Rapid Point-Implications for Rapid Point-of-Care Testingof-Care Testing• Change in HIV testing algorithm applies to clinical

labs only

• Confirmation of HIV infection cannot be made at the point-of-care using CLIA-waived tests

• Test providers should be aware of the new algorithm and with the types of supplemental tests that may be used to confirm preliminary positive results

FDA-Approved Rapid HIV Antibody Screening FDA-Approved Rapid HIV Antibody Screening TestsTests

Test Specimen Type

CLIA Category

Sensitivity Specificity Manufacturer

OraQuick ADVANCE Rapid HIV-

1/2 Antibody Test

Oral fluid Waived 99.3% 99.8% OraSure Technologies, Inc.

www.orasure.com

Whole Blood (f-stick,

venipunct)

Waived 99.6% 100%

Plasma Moderate Complexity

99.6% 99.9%

Uni-Gold Recombigen

HIV

Whole blood (f-stick,

venipunct)

Waived 100% 99.7% Trinity Biotech

ww.unigoldhiv.com

Serum & Plasma

Moderate Complexity

100% 99.8%

Reveal G-3 Rapid HIV-1

Antibody Test

Serum Moderate complexity

99.8% 99.1% MedMira, Inc.www.medmira.com

Plasma Moderate complexity

99.8% 98.6%

FDA-Approved Rapid HIV Antibody Screening FDA-Approved Rapid HIV Antibody Screening TestsTests

Test Specimen Type

CLIA Category*

Sensitivity(95% CI)

Specificity(95% CI)

Manufacturer

MultiSpotHIV-1/HIV-2

Rapid Test

Serum Moderate complexity

100% 99.93% BioRad Laboratorieswww.biorad.

comPlasma Moderate Complexity

100% 99.91%

Clearview HIV 1/2

STAT-PAK

Whole Blood (f-stick,

venipunct)

Waived 99.7% 99.9% Inverness Medical

Professional Diagnostics

www.invernessmedicalpd.com

Serum & Plasma

Non-waived 99.7% 99.9%

ClearviewCOMPLETE

HIV ½

Whole Blood (f-stick,

venipunct)

Waived 99.7% 99.90% Inverness Medical

Professional Diagnostics

www.invernessmedicalpd.com

Serum & Plasma

Non-waived 99.7% 99.9%

Sensitivity for Early HIV Infection of Rapid HIV Tests Sensitivity for Early HIV Infection of Rapid HIV Tests Compared with 3Compared with 3rdrd and 4 and 4thth Generation Assays Generation Assays J Clin Virol 2012;54:42-47J Clin Virol 2012;54:42-47

HIV Screening Assay No. Specimens Testing POS

Total No. Tested

Sensitivity for Early HIV Infection (%)

Architect HIV-1 Ag/Ab Combo 29 33 87.8Determine HIV-1 Ag/Ab Rapid Test (Alere, FDA-cleared 8/2013)

25 33 75.8

Genetic Systems HIV-1/2+O 19 33 57.5Multispot HIV-1/2 Rapid Test 11 33 33.3Clearview Complete HIV-1/2 8 27 29.6Unigold Recombingen HIV 8 33 24.2Clearview HIV-1/2 Stat-Pak 7 31 22.6Oroquick Advance HIV-1/2 7 32 21.9

Use of Fourth Generation IA’s and Use of Fourth Generation IA’s and Supplemental TestsSupplemental Tests• Improved HIV IA’s enhance ability to detect HIV infection earlier

• Acute infection, when substantial HIV transmission occurs• Specimens with reactive IA’s and negative supplemental test

results must undergo further testing to differentiate acute HIV infection from false-positive results

• Acute HIV infections detected with 3rd or 4th generation EIAs may be misclassified as HIV-negative by WB/IFA → • Adverse clinical outcomes for patients • Further HIV transmission within the community

• With FDA-clearance of Multispot as a supplemental test, labs can now adopt this algorithm

• Fast TAT enables delivery of same-day definitive test results• 3rd and 4th generation EIA’s → <1 hour • Multispot → 15 min

New HIV Testing Algorithm: New HIV Testing Algorithm: ConclusionsConclusions• Sensitive 3rd or 4th generation HIV-1/2 IA• If REACTIVE, a supplemental test (i.e., Multispot) is used to

differentiate HIV-1 and HIV-2 antibodies• If the supplemental test is REACTIVE for HIV-1 antibodies

• Confirmed HIV-1 Infection

• If the supplemental test is REACTIVE for HIV-2• Confirmed HIV-2 Infection

• If the supplemental test is discrepant with the initial IA result, a NAT test is recommended• Distinguish acute early infection from false-positive IA

Diagnosis Diagnosis of Syphilisof Syphilis

• Treponema pallidum cannot be cultured

• Primary syphilis diagnosed by direct detection methods (lesions)

• Darkfield microscopy

• Direct fluorescent antibody test for T. pallidum (DFA-TP)

• Polymerase Chain reaction

• Methods not widely available

• Direct detection methods can miss up to 30% of primary cases

• Most patients present without symptoms or signs of syphilis

• Healed early lesions

• Inapparent lesions

• Latent infections

• Syphilis is usually diagnosed by serologic tests

Lesions of Primary and Secondary Lesions of Primary and Secondary SyphilisSyphilis

Serologic Diagnosis of SyphilisSerologic Diagnosis of Syphilis

• Serologic diagnosis always requires detection of two types of antibodies

• Nontreponemal antibodies• Antibodies directed against lipoidal antigens

• Damaged host cells• Possibly from treponemes

• Treponemal antibodies• Antibodies directed against T. pallidum proteins

Serologic Serologic Diagnosis of Diagnosis of SyphilisSyphilis

• Nontreponemal tests• Rapid plasma reagin

(RPR) test• Venereal disease

research laboratory (VDRL) test

• Toluidine red unheated serum tests (TRUST)

• Treponemal tests• Fluorescent treponemal

antibody absorbed (FTA-ABS) test

• Treponema pallidum particle agglutination (TP-PA) test

Serologic Diagnosis of SyphilisSerologic Diagnosis of Syphilis

FTA-ABS

T. pallidum Particle Agglutination (TP-PA) /

T. pallidum HemAgglutination assays

(TP-HA)

Serologic Serologic Diagnosis of Diagnosis of SyphilisSyphilis

• Enzyme immunoassays• Trep-Chek (Phoenix

Biotech)• Trep-Sure (Phoenix

Biotech)• Chemiluminescence

immunoassays (CIAs)• Liaison• Architect

• Microbead immunoassays (MBIA)• BioPlex 2200

Syphilis IgG →→→• BioPlex 2200 IgM

Serologic Reactivity in SyphilisSerologic Reactivity in Syphilis

Traditional Testing Algorithm for Traditional Testing Algorithm for Diagnosis of SyphilisDiagnosis of Syphilis

Syphilis - Syphilis - Serologic Serologic Screening Screening Algorithms Algorithms – – Reverse Reverse Sequence Sequence

Syphilis Serology Syphilis Serology • Traditional algorithm

• Detects active infection• High rate of biological false-positives

• Confirmation with a treponemal test• Use of both tests results in a high PPV

• Can miss early primary and treated infections

• Reverse sequence algorithm• Detects early primary and treated infections that might be

missed with traditional screening• Non-treponemal test needed to detect active infection• Ideally, EIAs and CIAs should have perfect specificity

• False-positive results do occur• Varies by risk group being tested

Syphilis Immunoassays - TimelineSyphilis Immunoassays - Timeline

1980’s EIA is FDA-cleared for use as a confirmatory test and in blood bank screening

2000 UK Public Health Lab Guidelines says EIA is an “appropriate alternative” to VDRL/RPR and TPHA

2001 EIA is FDA-cleared for clinical diagnostic use

2008 EU Guidelines: EIA/TPPA recommended for screening; VDRL and RPR no longer recommended

2009 CDC-APHL Report: Presents algorithm for screening with treponemal EIA

Sensitivity and Specificity of Sensitivity and Specificity of Serologic Tests for SyphilisSerologic Tests for Syphilis

Why Switch to Why Switch to EIA/CIA?EIA/CIA?

• Automated (high throughput)• 180 tests per hour

• Low cost in high-volume settings

• Less lab occupational hazards• No manual pipetting

• No false-negatives due to prozone reaction

Why Switch to Why Switch to EIA/CIA?EIA/CIA?

• Objective results• Some EIA/CIA test

detect IgM antibodies• BioPlex 2200

assay• Potentially useful

for diagnosis of early syphilis

Challenges and Limitations of Challenges and Limitations of EIA/CIAEIA/CIA• Cannot distinguish between active disease and old

disease (treated/untreated)• Studies to compare test performance with other

serologic tests are lacking• Studies evaluating performance of EIA/CIA to detect

IgM antibodies are lacking but ongoing• Confusion regarding management of patients with

discrepant serology• Positive EIA/CIA result and a negative RPR

Reasons for Discordant Test Reasons for Discordant Test Results: EIA/CIA+ → RPR-Results: EIA/CIA+ → RPR-• False-positive EIA/CIA

• Very sensitive• Lower specificity

• Treated syphilis• Treponemal antibodies are detected by sensitive

EIAs and CIAs• Sero-reversion of non-treponemal antibodies

• Early primary syphilis• Treponemal antibody titer rises before non-

treponemal antibody titer

Interpretation of Serologic Tests Interpretation of Serologic Tests for Syphilisfor Syphilis• Reactive results in both treponemal and RPR tests →

• Untreated syphilis (unless ruled out by treatment history) • Persons treated in the past are considered to have a new

infection if quantitative RPR testing reveals a four-fold or greater increase in titer

• Reactive result in the treponemal test but non-reactive in the RPR test →• Those with a history of previous treatment require no further

management• For those without a history of treatment, a second, different

treponemal test should be performed (i.e., TPPA)• If the second treponemal test is non-reactive, no further

evaluation or treatment is indicated, or perform a third treponemal test to resolve the discrepancy

Reverse Sequence Algorithm Reverse Sequence Algorithm for Syphilisfor Syphilis• Composite

results of reverse sequence algorithm for initial screening

ConclusionsConclusions• EIA/CIA have high sensitivity but lower specificity• All reactive EIA/CIA must be reflexed to a quantitative RPR

• Confirms reactive EIA/CIA• Detects active infection

• Although test performance varies by prevalence of syphilis in the population, all discordant specimens (i.e., EIA+/RPR-) must be confirmed with a confirmatory treponemal test

• Confirmatory treponemal test must have at least equivalent sensitivity and a higher specificity compared to the screening treponemal tests (EIA/CIA)• TP-PA recommended• FTA-ABS not recommended

Nucleic Acid Amplification Tests for Nucleic Acid Amplification Tests for N. N. gonorrhoeaegonorrhoeae and and Chlamydia trachomatisChlamydia trachomatis

Test Manufacturer Method of Detection

FDA -Cleared Not FDA-Cleared

Amplicor Roche Molecular Diagnostics

PCR Female endocervical;Male urethral;Male and female FV urine;Self-collected vaginal swabs

Extragenital sites;Children (any site)

Probe-Tec Becton-Dickinson

Strand displacement amplification

Female endocervical;Male urethral;Male and female FV urine;Self-collected vaginal swabs

Extragenital sites;Children (any site)

APTIMA Hologic/Gen-Probe

Transcription-mediated amplification

Female endocervical;Male urethral;Male and female FV urine;Self-collected vaginal swabs

Extragenital sites;Children (any site)

Real-Time m2000

Abbott Molecular Diagnostics

Real-time PCR

Female endocervical;Male urethral;Male and female FV urine;Self-collected vaginal swabs

Extragenital sites;Children (any site)

NAATS for GC/CTNAATS for GC/CT• Advantages over culture-based methods

• Greater sensitivity• Use of non-invasive specimens

• Limitations (especially for GC)• Genetic variation/recombination can affect gene targets for

amplification, leading to potentially false-negative results• Horizontal interspecies exchange of genetic material

between Neisseria species may lead to false-positive results when commensal Neisseria acquire gonococcal sequences and vice versa

• PCR and SDA have both demonstrated cross-reactivity with other Neisseria species

Gene Targets and Cross-Gene Targets and Cross-Reactivity for Gonococcal NAATsReactivity for Gonococcal NAATsTest Amplification Target Positive Reactivity with other Neisseria

Species

Roche Amplicor

Cytosine DNA methyltransferase gene (single copy)

N. meningitidis, N. lactamica, N. subflava/sicca, N. cinerea, N. flavescens, N. polysaccharea, M. catarrhalis

Gen-Probe APTIMA 2

16S subunit of ribosomal RNA gene (multicopy)

None reported

BD Probe-Tec

Multi-copy pilin gene inverting protein homolog

N. meningitidis, N. lactamica, N. cinerea, N. mucosa, N. flavescens,

Abbott Real-Time PCR

Opa gene (multicopy) None reported

NAATS for Detection f NAATS for Detection f N. gonorrhoeae N. gonorrhoeae in in Oropharyngeal and Rectal SitesOropharyngeal and Rectal Sites

• McNally et al, CID 47:e25-e27, 2008• SDA had low positive predictive value for oral (30.4%) and

rectal (73.7%) specimens in an MSM population

• Schachter et al, STD 35:637-642, 2008• Sensitivities of NAATS (PCR, SDA, TMA) were better than

culture for detection of oral/rectal GC in MSM • Specificity of PCR 78.9% for oral swabs• Specificity of SDA and TMA ≥99.4 for oral/rectal sites

• Bachmann et al, JCM 42:902-907, 2009• PCR had specificity of 73% compared to 96.3% for SDA and

98.6% for TMA for oropharyngeal GC infection in population with acknowledged oropharyngeal sexual contacts

Use of NAATS for Oropharyngeal GCUse of NAATS for Oropharyngeal GCBachmann et al, JCM 42:902-907, 2009Bachmann et al, JCM 42:902-907, 2009

• Evaluated PCR (Roche), TMA (Gen-Probe) and SDA (BD Probe-Tec)

• NAATS were compared with culture• Males and females who acknowledged oral sexual

contacts in the previous two months recruited from 3 clinics in Birmingham, AL

• Data evaluated using a “rotating gold standard”• Any positive results by two or three of the three

tests that excluded the test being evaluated• 961 evaluable test sets obtained

Use of NAATS for Oropharyngeal GCUse of NAATS for Oropharyngeal GCBachmann et al, JCM 42:902-907, 2009Bachmann et al, JCM 42:902-907, 2009

TEST Sensitivity Specificity

Positive results by two of three comparator tests

Culture 50% 99.4%

PCR 80.3% 73.0%

SDA 93.2% 96.3%

TMA 83.6% 98.5%

Positive results by three of three comparator tests

Culture 65.4% 99.0%

PCR 91.9% 71.8%

SDA 97.1% 94.2%

TMA 100% 96.2%

Use of NAATS for Oropharyngeal/Rectal GC/CT Use of NAATS for Oropharyngeal/Rectal GC/CT Schachter et al STD 35:637-642, 2008Schachter et al STD 35:637-642, 2008

• Specimens from 1110 MSM• Evaluated PCR, SDA, and TMA compared with

culture• True-positive GC

• Culture positive, OR• TMA/PCR positive or TMA/SDA positive, OR• A single positive NAAT confirmed by an alternate target

NAAT

• True-positive CT• Culture positive, OR • Two positive NAATS, OR• A single positive NAAT confirmed by an alternate target

NAAT

Use of NAATS for Oropharyngeal/Rectal GC/CTUse of NAATS for Oropharyngeal/Rectal GC/CTSchachter et al STD 35:637-642, 2008Schachter et al STD 35:637-642, 2008

• Based on initial findings with 205 MSM specimens, PCR had a 78.9% GC specificity with oropharyngeal swabs, so PCR testing was discontinued

• Oropharyngeal GC infections (89 infections detected)• Sensitivity of Culture →→ 41%• Sensitivity of SDA →→→ 72%• Sensitivity of TMA →→→ 93%• Specificity of SDA/TMA → ≥99.4%

• Rectal GC infections (88 infections detected)• Sensitivity of Culture →→ 43%• Sensitivity of SDA →→→ 78%• Sensitivity of TMA →→→ 93%• Specificity of SDA/TMA → ≥99.4%

Use of NAATS for Oropharyngeal/Rectal GC/CTUse of NAATS for Oropharyngeal/Rectal GC/CTSchachter et al STD 35:637-642, 2008Schachter et al STD 35:637-642, 2008

• Oropharyngel CT infections (9 infections detected)• Sensitivity of culture →→→ 44%• Sensitivity of SDA →→→→ 78%• Sensitivity of TMA →→→→ 100%• Specificity of SDA/TMA →→≥99.4%

• Rectal CT infections (68 infections detected)• Sensitivity of culture →→→ 27%• Sensitivity of SDA →→→→ 63%• Sensitivity of TMA →→→→ 93%• Specificity of SDA/TMA →→≥99.4%

Use of NAATS in Children – 2010 Use of NAATS in Children – 2010 CDC STD Treatment GuidelinesCDC STD Treatment Guidelines• Recommend initial culture of the pharynx, anal canal, and genital

tracts for GC for both boys and girls• Recommend CT cultures from the anal canal in boys and girls and

from the vagina in girls

• NAATS can be used for detection of GC and CT in vaginal swabs and urine from girls being evaluated for suspected sexual abuse

• NAATS not recommended for use in boys or for extragenital infections in children, as there are no supporting data

• For cases of suspected sexual abuse, confirmatory testing by a second NAAT should be performed

• Laboratories should use newer “second generation” NAATs with the highest sensitivity possible, preferably with a different target

• Specimens should be retained for further testing

Diagnostic Issues Raised by Using Diagnostic Issues Raised by Using NAATS for Non-FDA-Cleared Sites NAATS for Non-FDA-Cleared Sites

• Validity of NAAT results may be legally challenged• Address concerns, in part, by collection of

concurrent/follow-up cultures• Reported results should include the caveat that the

site of the specimen is not FDA-cleared (off-label)• Under U.S. law, laboratories may offer testing for

extra-genital gonococcal or chlamydial infection if “internal validation of the method by a verification study” is performed

Prepubertal Prepubertal Non-Acute Non-Acute Sexual AbuseSexual Abuse

• Acquire site-specific specimens from: • Asymptomatic children

who disclose contact• Asymptomatic children

with known/highly suspected contact with an infected individual

• Those with an allegation of abuse and symptoms of infection

• All positive NAATs confirmed by a second NAAT and/or culture

Pubertal Non-Pubertal Non-Acute Sexual Acute Sexual AbuseAbuse

• American Academy of Pediatrics recommends use of urine-based and vaginal swab NAATS (oral/anal NAAT testing not addressed)

• Site-specific specimens to detect STI’s by NAAT obtained at first visit

• Repeated two weeks later

• All positive tests confirmed by an additional NAAT and/or culture

Thank You!Thank You!