hiv testing technologies - caiglobal.org...bio-rad gs hiv combo ag/ab eia •microwell plate eia...
TRANSCRIPT
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HIV Testing Technologies
Barbara G. Werner, PhD Infectious Disease Consultant
MA Department of Pubic Health December 5, 2013
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Disclosures
I, Barbara G. Werner, PhD, have no relevant financial, professional or personal relationship to disclose.
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Learning Objectives
• Discuss current HIV diagnostic tests
• Identify limitations of the current HIV diagnostic algorithm
• Describe revised HIV diagnostic algorithm
• Describe the use of the new algorithm to identify acute HIV infections
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Acknowledgement
Thanks to Bernard M. Branson, MD for sharing a number of slides used in this presentation.
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1st and 2nd Generation EIA Plasma/serum (1 h/37o C)
Plasma/serum
Color
reagent
Antigen
1st - Viral lysate
2nd – Recombinant proteins or
synthetic peptides
IgG HIV antibody
enzyme
anti-human IgG
Detects HIV
IgG
Enzyme-
detection
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Available 1st & 2nd Generation Assays
• HIV-1 Western blot
• HIV-1 IFA
• HIV-1 EIA
• Six rapid HIV antibody tests
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EIA
Western blot
IFA
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Uni-Gold Recombigen
OraQuick Advance Clearview Complete
Clearview Stat Pak INSTI
CLIA Waived POC Rapid Tests
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Multispot HIV-1/HIV-2 Rapid Test
• CLIA moderate complexity with serum, plasma
• Perform test in 15 minutes
• Shelf life: 1 year refrigerated, 3 months room temperature
• FDA-approved for use in algorithm March 28, 2013
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FDA-approved HIV-1/HIV-2 Antibody Differentiation Assay
Peptide HIV-2
Recombinant HIV-1
Peptide HIV-1
Reactive Control
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3rd Generation “Sandwich” EIA
Antigen:
Recombinant proteins or
synthetic peptides Enzyme-
detection
Color
reagent
Plasma/serum HIV antibody
enzyme
HIV antigen
Detects HIV
IgM or IgG
IgG IgM
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Available 3rd Generation Assays
• One HIV-1/HIV-2 EIA
– GS HIV-1 HIV-2 Plus O
• Two HIV-1/HIV-2 CIAs
– Advia Centaur 1/O/2
– Ortho Vitros HIV1+2
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4th Generation Combo Assay
Color
reagent
Enzyme-
detection
HIV antigen
p24 antibody
Plasma/serum HIV antibodies
p24 antigen
HIV antigen
p24 antibody
Detects IgM
or IgG
antibody or
p24 antigen
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Available 4th Generation Assays
• One HIV-1/HIV-2 antigen/antibody EIA
• One HIV-1/HIV-2 antigen/antibody CIA
• And most recently, an HIV-1/HIV-2 antigen/antibody rapid test which distinguishes antigen from antibody
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• Chemiluminescent immunoassay
• Detects p24 antigen and HIV antibody
• Time to result: 29 minutes
• FDA-approved June 22, 2010
Abbott Architect 4th Generation Ag/Ab
Combo Assay
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Bio-Rad GS HIV Combo Ag/Ab EIA
• Microwell plate EIA
• 3rd generation Ab format:
- HIV-1: gp160
- HIV-2: gp36
- Group O
• p24 antigen
• FDA-approved July 25, 2011
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Determine Combo Rapid HIV 1/2 Ag/Ab Test
• CLIA moderate complexity
• Distinguishes Ag from Ab
• Whole blood, serum plasma
• FDA-approved August 2013
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Antibody
Antigen
Control
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APTIMA Qualitative HIV-1 RNA Assay Aid to HIV-1 diagnosis
Diagnosis of acute HIV-1 infection in antibody-negative persons
Confirmation of HIV-1 infection in antibody-positive persons when RNA is detected
FDA-approved July 2006
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Laboratory-based Algorithm
• From late 1980s
• Initial screening test, generally EIA
• Supplemental test, WB or IFA, performed following repeatedly reactive EIA
• Results reported as Positive, Negative, or Indeterminate
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Limitations of the Current Algorithm
• Antibody tests do not detect infection in ~10% of infected persons at highest risk of transmission---miss acute infections
• Western blot confirmation is less sensitive during early infection than many widely used screening tests---indeterminate or false negative results
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Further Limitation
• HIV-2 infections may be miss-classified as HIV-1 on Western blot – 54/58 (93%) HIV-2 patients tested had positive
HIV-1 WB (NYC)*
– 97/163 (60%) HIV-2 cases reported had positive HIV -1 WB (CDC)**
• HIV-2 remains uncommon in U.S., but – Does not respond to NNRTIs, some PIs (first line
therapy) – Undetectable by HIV-1 viral load tests
*Torian et al.,CID 2010; **MMWR July 2011
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Sequence of Test Positivity Relative to WB (serum)
166 specimens, 17 Seroconverters - 50 % Positive Cumulative Frequency
Modified from Masciotra et al, J Clin Virol 2011
Days before WB positive
WB
po
sit
ive
AP
TIM
A (
-26
)
GS
1/2
+O
(-
12
)
Mu
lti-
Sp
ot
(-7
) R
eve
al G
3, (-
6)
Vir
on
os
tka
(+2
)
Ora
Qu
ick
(-1
)
Un
igo
ld (
-2)
25
20
10
5
0
15
Arc
hit
ec
t A
g/A
b C
om
bo
(-
20
)
CO
MP
LE
TE
HIV
-1/2
(-
5)
H
IV-1
/2 S
TA
T-P
AK
(-
5)
Ad
via
(-
14
) V
itro
s (-
13
)
INS
TI (
-9)
Bio
-Ra
d A
g/A
b C
om
bo
(-
19
)
and Owen et al, J Clin Micro 2008
Avio
q
De
term
ine
Ag
/Ab
Co
mb
o (-
15
)
DP
P (
-8)
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HIV RNA (plasma)
HIV Antibody
0 10 20 30 40 50 60 70 80 9
0
100
HIV p24 Ag
22
1st
gen
2nd
gen
3rd
gen
Days
Modified after Busch et al. Am J Med. 1997
Infection 4th
gen
Acute HIV Infection
HIV Infection and Laboratory Markers
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New HIV Diagnostic Algorithm for serum and plasma
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Some Alternatives and Limitatons 1. If 3rd gen HIV-1/2 IA as initial test: perform subsequent
testing specified in the algorithm. Will miss some acute infections
2. If alternative 2nd Ab test is used (e.g., WB or IFA): If
negative or indeterminate, perform HIV-1 NAT; if HIV-1 NAT is negative, perform Ab IA for HIV-2
Might miss-classify HIV-2 infections; requires more tests; increases TAT
3. HIV-1 NAT as 2nd test: if positive, HIV-1 infection; if
negative, perform HIV-1/HIV-2 Ab differentiation assay. Won’t distinguish acute from established infections; increases TAT and costs
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HIV
RN
A in
Sem
en
(Lo
g 10 c
op
ies/
ml)
2
3
4
5
1/1000 - 1/10,000
1/500 - 1/2000
1/100- 1/1000
1/30- 1/200
Increased Risk of Sexual Transmission of
HIV
Cohen & Pilcher, J Infect Dis. 2005
Virus 75-750 times more infectious
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Detecting Acute HIV Infections • Sensitivity among frequently-tested MSM in
Seattle
– 192 infected with HIV
• 23 (12%) detected only by RNA
– (15/16 tested detected by Ag/Ab immunoassay)
• 169 (88%) detected by serum Ab immunoassay
• 153 (80%) detected by oral fluid rapid test
- Stekler et al, Clin Inf Dis 2009
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Phoenix ED Screening July 2011 through February 2013
• 4th gen screening of patients who had blood drawn – 15% of patients declined testing
– 13,014 patients tested
– 37 (0.3%) new HIV infections • 12 (32.4%) had Acute HIV Infection (antibody negative)
• Median viral load: – Patients with acute infections: 6 million
– Patients with established infections: 25,000
-MMWR June 21, 2013
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RNA vs. 4th Generation Ag/Ab Assay
• RNA+/ 3rd gen-negative specimens detected by 4th generation EIA: – 38 of 46 (83%) – Australia* – 10 of 14 (71%) – CDC AHI study** – 51 of 61 (84%) – CDC panel***
– 4 days after RNA – 9 seroconversion panels***
* Cunningham P, HIV Diagnostics Conf 2007 ** Patel P, CROI 2009 *** Owen M, CROI 2009
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4th generation HIV-1/2 immunoassay
HIV-1/HIV-2 antibody differentiation immunoassay
(-) (+)
HIV-1 (+)
HIV-2 (-)
HIV-1 antibodies
detected
HIV-1 (-)
HIV-2 (+)
HIV-2 antibodies
detected
HIV-1 (-) or indeterminate
HIV-2 (-)
NAT
NAT (+) Acute HIV-1 infection
NAT (-) Negative for HIV-1
Negative for HIV-1 and HIV-2
antibodies and p24 Ag
HIV-1 (+)
HIV-2 (+)
HIV antibodies
detected*
*Additional testing required to rule out dual infection
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Use of the New Algorithm MA State PHL- 1st year results with 4th gen
• 7984 sera tested (BioRad “Combo”)
• 258 (3.2%) positive for HIV-1
• 1 positive for HIV-2
• 16 4th gen EIA RR and MS negative
– 8 had detectable HIV RNA, i.e., acute infections
– 8 had no detectable HIV RNA, i.e., likely false positive on initial screen
Goodhue etal. 2013 JClinVirol
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Acute HIV Infections in MA Retrospective analysis of 8 specimens from acutely infected persons:
5 were negative on a 3rd generation test and would have been reported as negative
3 reactive specimens were run on Western Blot
None were positive
2 were indeterminate and 1 was negative
At best, confirmation of infection would have been delayed until a follow up specimen was obtained.
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Acute infections in MA
• 8 individuals were given accurate, timely information about their infection status and linked to clinical care.
With a collaborative effort of lab, program, clinical providers, DIS and others, transmission can be impacted if acutely infected can be identified
Contacts were identified
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Characteristics of 8 Persons with Acute Infection
• Substantial sexual risk, primarily MSM
• Certain vulnerabilities, such as homelessness, limited English proficiency, recent immigration
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Resources • Criteria for Laboratory Testing and Diagnosis of Human
Immunodeficiency Virus Infection. Approved guideline. CLSI document M53-A. Wayne, PA: Clinical and Laboratory
Standards Institute: 2011. • Update on HIV Diagnostic Testing Algorithms. Journal of Clinical
Virology – open access 2011 and 2013 http://www.journalofclinicalvirology.com/supplements# • CDC/APHL Document. Updated Recommendations for Laboratory
Testing for Diagnosis of HIV Infection. To be published in 2014. • CDC and APHL and HIVtestingconference websites