HIV Diagnosis,
Acute Infection and
Superinfection
Don Kurtyka, ARNP, MS, MBA
University of South Florida College of Medicine
Tampa General Hospital
Hillsborough County Health Department
Objectives
 Discuss the diagnosis of HIV and available
tests
 Describe the approach to the diagnosis of
acute retroviral syndrome
 Debate the advantages and disadvantages
of early treatment of acute HIV infection
 Discuss the evidence for the possibility of
superinfection / reinfection and the
implications for patient education and
management
Anonymous vs Confidential
 Anonymous
 Identifying information not provided
 Results not linked to identifying information
 Allows reporting of HIV infection without breaching
confidentiality
 Disadvantage: may not be able to locate clients for test
results
 Confidential
 Clients linked to test result by identifying information
 Results remain confidential
 Informed consent
Pre-Test Counseling
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Goal: reduce HIV acquisition and transmission
Accurate and current information about HIV
Obtain informed consent
Transmission and acquisition
HIV test info: risk, benefits, meaning of potential
test results
 Assessment of individuals risks and appropriate
risk reduction activities
 Capacity to comprehend HIV testing and consent
Post-Test Counseling
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Accurate and current information about HIV
Local resources
Risk reduction education
Referrals for ongoing care and support
Healthy living strategies
Meaning of test results and state reporting
guidelines
 Mental health support / counseling
Diagnosis of HIV Infection
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Viral antibodies
Viral antigens
Viral RNA/DNA
Culture
Lancet, 1996; 348: 176.
Enzyme Immunoassay
Enzyme-Linked Immunosorbent Assay
(EIA, ELISA)
 Primary HIV antibody screening test
 Serum plasma, dried blood spots, oral fluids,
urine
 HIV-1/2, HIV-1, HIV-2
 High degree sensitivity and specificity
 Repeatedly reactive: confirmatory testing
Negative Antibody Test Results
 HIV negative
 Recent infection: too early for seroconversion
 CDC: follow-up testing at 6 weeks, 12 weeks,
6 months
Confirmation Process
 Non-negative screenings should be
confirmed
 Western Blot (WB)
 Immunofluorescent Antibody Assay (IFA)
 Higher specificity than EIA
 Interpretation can be subjective
Predictive Value: HIV Ab Tests
 Depends on the prevalence of HIV infection
in the population
 Low HIV prevalence: predictive value of a
positive test is low
 HIV Ab testing of low prevalence populations
likely to produce more false-positive than
true-positive results
Window Period
 Time delay from infection to positive EIA
 Average: 10-22 days
 Most seroconvert within six months
Am J Med 2000; 109
HIV-1 vs HIV-2
 HIV-1: Most cases
 Group M: predominant strain world-wide
 Subtypes (clades): A to K, N, O
 Clade B
 US and Europe
 98% of HIV-1 in US
 Most non-B subtypes were acquired outside US
 Clade C: Southeast Asia
 N (“new”): 1998
 Group O: West Africa
 Recombination between viruses of different clades
becoming more common
Predominant HIV-1 Subtypes
 A: West/East/Central Africa, East Europe,
Mideast
 B: North America, Europe, Mideast, East
Asia, Latin America
 C: South Africa, South Asia, Ethiopia
 D: East Africa
 E: Southeast Asia
JAIDS 2002; 29:184
HIV-2
 Primarily found in West Africa
 Causes immune deficiency due to depletion of CD4
cells
 5-8 fold less efficient transmission compared to
HIV-1
 Associated with lower viral load
 Slower rate of CD4 decline and clinical progression
 Negative Ab tests in 20-30% depending on EIA
assay
 WB: not well standardized nor FDA approved
Bartlett, JG 2003: Medical Management of HIV Infection, p5.
Testing Recommendations: HIV-2
 Natives of endemic areas
 Needle-sharing and sex partners of persons
from endemic areas
 Sex or needle-sharing partners of persons
with known HIV-2 infection
 Transfusion or non-sterile injection recipients
in endemic areas
 Children of HIV-2 infected women
Other
West Africa
HIV-2 Endemic Areas
Benin
Burkina Faso
Cape Verde
Cote d’Ivoire
Gambia
Ghana
Guinea Guinea-Bissau
Liberia
Mali
Mauritania
Niger
Nigeria
Sao Tome
Senegal
Sierrra Leone
Togo
Mozambique
Angola
Confirmation Process: WB
 Detects antibodies to HIV-1 proteins
 Core: p17, p24, p55
 Polymerase: p31, p51, p66
 Envelope: gp41, gp120, gp160
 Negative: no bands
 Positive:
 Reactivity to gp41 + gp120/160 or
 Reactivity to p24+gp120/160
 Indeterminate:
 EIA repeatedly reactive
 Presence of any band pattern not meeting criteria for
positive results
False Negative Results
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High-prevalence population: 0.3%
Low-prevalence: <0.001%
Usually due to testing during window period
Rare patients seroconvert in late-stage
disease
 Technical or clerical error
 Type N or O
 HIV-2
False Positive Test Results
 Much less common than in earlier times
 Frequency: 0.0004% to 0.0007%
 Causes
 Autoantibodies (single case, Lupus, ESRD)
 HIV vaccines
 EIA+: 68%
 WB+: 0-44%
 Technical / clerical error
NEJM 1988;319:961
Ann Intern Med 1989;110:617
Indeterminate Results
 4-20% of WB assays with positive bands
 Testing during seroconversion
 p24 usually appears first
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Late stage HIV: loss of core antibody
HIV vaccine recipients
Technical / clerical error
Infection with O strain or HIV-2
Indeterminate Results (continued)
 Cross-reacting nonspecific antibodies
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Collagen-vascular disease
Autoimmune disease
Pregnancy
Organ transplantation
Lymphoma, other malignancies
Liver disease
Multiple sclerosis
Recent immunization
Indeterminate Results
 Evaluate HIV risk
 Low risk: almost never infected with HIV-1 or
HIV-2
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Repeat testing: often continued indeterminate
Cause: frequently not established
HIV unlikely
Follow-up serology in 3 months
 Seroconversion: usually WB+ in 1 month
 Repeat testing at 1, 2, 6 months
 Counseling to reduce potential transmission
Frequency of HIV Testing
 High risk behavior: every 6-12 months
 Annual seroconversion
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General population: 0.02%
Military recruits: 0.04%
MSM: 0.5 - 2%
IDU in high prevalence area: 0.7-6%
Alternative Testing
 Home test kits
 Rapid Testing
 Alternative body fluids
 Saliva
 Urine
 Vaginal secretions
 Viral detection
Home Testing
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Home specimen collection
Self-dried blood spot obtained with lancet
Anonymous coding
Mail/courier to testing facility
Double EIA and confirmatory IFA/WB
Sensitivity/Specificity: ~100%
Results relayed to user by telephone after user
initiates request
 Negative: prerecorded message
 Positive: live conversation and counseling
Rapid HIV Antibody Detection
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Results in 15-20 minutes
Occupational exposure
Women in labor with unknown HIV status
Clients unlikely to return for visits
Outreach
ERs
Rapid HIV Antibody Detection
 OraQuick HIV-1 Antibody Test (OraSure)
 Results read by provider in 20 minutes
 Sensitivity: 99.6% / Specificity: 100%
 $20-30
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Testing initially delayed due to CLIA requirements
Fingerstick sample of blood
Negative test: definitive
Positive test: needs standard serology confirmation
Not recommended for HIV-2 screening
Rapid HIV Antibody Detection
 Single Use Diagnostic System (SUDS) HIV-1
Test
 Venipuncture
 Results: 15-30 minutes
 Confirmatory WB required
 Double Check (Organies)
Type N, Type O, HIV-2
 EIA may fail to detect O subtype
 N group: causes false-negative EIA but may
be WB positive
 HIV-2: false negative EIA in 20-30%
 Consider specific HIV-2 testing
P24 Antigen
 Part of blood bank algorithms since 1996
 Uncommon in clinical practice
 Detects free, non-complex HIV antigens in
peripheral blood
1 mil
HIV
RNA
100,000
+
_
10,000
Ab
P24 +
1,000
100
Exposure
Symptoms
10
0
20
30
Days
40
50
HIV-1 Antibodies
HIV RNA
Typical Course of Primary
HIV
Rapid Test Results
 Reactive (preliminary positive) rapid test
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Screening test is positive
Preliminary result
Confirmatory testing required
Precautions to avoid viral transmission
 Negative rapid test
 No recent exposure: definitive negative
 Possible recent exposure:
 Recommend re-test
 Counseling to prevent transmission
OraQuick: Florida DOH
 6 Month Pilot Studies
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Hillsborough CHD
Duval County Jail
Orlando CBO for substance abuse
Miami: 2 sites
Key West: only anonymous site
Saliva Testing: OraSure
 EIA and WB to detect IgG
 Specimen collection device, antibody screen, WB
confirmation
 Cost: ~$25
 Specially treated pad placed between lower cheek
and gum for 2 minutes
 Vial sent to lab for processing
 Sensitivity and specificity comparable to standard
serologic testing (~99.5%)
 Advantages: ease of collection; low cost; improved
patient acceptance
 Disadvantage: client must return for results
Urine Testing
 Calypte HIV-1 Urine EIA
 Positive results require standard serologic
confirmation
 Sensitivity: 99%; Specificity: 94%
 Cost: ~$4
Vaginal Secretions
 IgG EIA
 CDC: recommended for rape victims
 Semen contains HIV IgG Ab
Indications for HIV Viral
Detection
 Confusing / indeterminate serologic test
results
 Acute retroviral infection
 Neonatal infection
 Window period following exposure
 Not FDA approved for diagnosis of HIV
 Expensive
Viral Detection
 p24 Antigen
 HIV-1 DNA PCR
 Most sensitive: able to detect 1-10 copies of
proviral DNA
 S/S: 99% / 98%
 HIV-1 RNA (RT-PCR, bDNA)
 S/S: 95-98%
 Viral culture of PBMC: expensive, labor
intensive, reliability variable
Viral Detection: HIV-2
 bDNA proficient at quantitation of many nonclade B viruses
 Amplicor version 1.5 designed to detect other
clades
National Recommendations
For HIV Testing of
Pregnant Women
 USPHS Recommendations for HIV Screening of
Pregnant Women (4-22-03)
 Universal testing for all pregnant women as a routine
part of prenatal care using an “opt out” approach
 Labor and Delivery: routine rapid testing if HIV status
unknown
 Postnatal: rapid testing for all infants whose mother’s
status is unknown
 Regulations, laws, and policies about HIV
screening of pregnant women vary from state to
state
Acute HIV Infection
Acute HIV Infection
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Transient symptomatic illness in 40-90%
Usually mild but can be severe
2-6 weeks after infection
Often not recognized by primary care clinicians
 Symptoms non-specific
 Often resembles influenza, mononucleosis
 “Cold symptoms” absent
 Can be asymptomatic
 Duration: 14 days
DHHS Guidelines July 14, 2003
HIV testing
• CDC statistics – illness is still the most common reason for testing
• ER study (inner-city Baltimore)
– Prevalence 11.3%, with majority being AA males, followed by AA women3
• Think HIV – urgent care sites in high-prevalence regions of MA4
– HIV prevalence was 2.2%, higher than state-funded testing programs
– Testing offered to all, not just those perceived as high-risk
– Intense focus on linkage to care: 88% return rate for those HIV+
• Failure to return for results:
– 508 subjects, 55% failed to return for results2
• Opt-out (actively refusing testing) leads to greater acceptance than opt-in
(actively agreeing to testing): 85-98% vs 25-83%1
1. Mbori-Ngacha D. 10th CROI, Boston 2003; #47; 2. Hightow LB, et al. ibid; #918; 3. Henson C, et al. ibid; #38;
4. Walensky RP, et al. ibid; #39
14
Acute HIV Infection
Fever
LAD
Pharyngitis
Rash
96%
74%
70%
70%
Headache
N/V
HSM
Wt loss
32%
27%
14%
13%
Myalgia/arthralgia
Diarrhea
54%
32%
Thrush
Neuro Sx
12%
12%
Neuro: meningoencepalitis or aseptic meningitis; peripheral neuropathy or radiculopathy;
facial palsy, Guillain-Barre syndrome; brachial neuritis; cognitive impairment or
psychosis
CDC 2002
Rash in Acute HIV Infection
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Trunk, face, extremities
Palms and soles rarely involved
5-10 mm diameter
Erythematous, nonpruritic, painless
Laboratory Findings
Acute HIV Infection
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Lymphopenia  lymphocytosis
Atypical lymphocytes
Transient CD4 decline
VL: 100,000 – 1,000,000
Diagnosis of Acute HIV Infection
 Recognition of clinical symptoms
 No true constellation of signs/sympoms
 Presence of any symptom(s)
 History of activity associated with HIV risk
 Detectable plasma HIV RNA
 Highly sensitive
 False positive possible
 Detectable p24 Antigen
 Less sensitive
 False positive rare
Acute HIV Infection
 High virus levels (105-106 copies/mL)
 2-9% of HIV-negative have false positive
results
 Usually associated with low RNA titers <10,000
 VL in new infections
 Correlates with rate of CD4 decline
 Prognostic indicator in early disease
Potential Benefits: Early Intervention
 Decrease the severity of acute disease
 Alter initial viral set point  alter disease
progression rate
 Suppress viral replication  reduce rate of viral
mutation
 Preserve HIV-specific immune responses
 May permit future discontinuation of therapy with
sustained viral control
 Reduce risk for viral transmission
 May minimize viral evolution and development of
viral diversity
DHHS Guidelines July 14, 2003
Potential Risks: Early Intervention
 Decreased QOL
 Medication side effects
 Drug toxicities
 Dosing constraints
 Drug resistance if viral suppression
inadequate
 Need for indefinite continuing therapy
 Expensive
 Potential for transmission of resistant virus
DHHS Guidelines July 14, 2003
Potential Risks: Early Intervention
 Long term clinical outcome benefit has not
been documented
 Additional studies are needed to delineate
the role of ARV therapy during the primary
infection period
DHHS Guidelines July 14, 2003
Treatment: Acute HIV Infection
 Weigh potential benefits against potential
risks
 “Certain authorities endorse treatment of
acute HIV infection on the basis of the
theoretical rationale and limited but
supportive clinical trial data”
DHHS Guidelines July 14, 2003
Treatment: Acute HIV Infection
 Experienced clinicians recommend consideration of
therapy for patients among whom seroconversion
has occurred within the previous 6 months
 “Although the initial burst of viremia among infected
adults usually resolves in 2 months, treatment
during the 2 to 6-month period after infection is
based on the probability that virus replication in
lymphoid tissue is still not maximally contained by
the immune system during this time”
DHHS Guidelines July 14, 2003
Detuned Antibody Testing
 Less sensitive ELISA test
 May help distinguish between recent seroconverters and
those with long-standing HIV infection
 Current ELISAs can detect relatively low levels of Ab
 HIV Ab levels increase over first few months
 Recent infection: standard ELISA positive
 Detuned assay: negative
 Able to diagnose individuals who have already
seroconverted on a standard ELISA but are still early in
infection
HIV Superinfection
HIV Super-Infection
 Coinfection with a second strain of HIV
during the course of established HIV-1
infection (Jost, NEJM 347:10, 2002)
 Known to be theoretically possible
 Little direct evidence to support concept
HIV Superinfection
 2000: LTNP (patient A) – unprotected
intercourse with ARV-experienced male with
progressive HIV disease (patient B)
 Patient A experienced rapid disease
progression
 Virus harbored original strain and drugresistant strain from patient B
Angel, J. CROI 2000, Abs LB2
HIV Superinfection
 Established infection with HIV-1, subtype AE
 Well-controlled viremia on HAART; unable to
remain on ART due to liver toxicity
 Sexual exposure to type B in Brazil
 Unprecedented rise in viral load and rapid
CD4 depletion
 Mixture of B and AE identified
 Rapid emergence of type AE
Jost, S. NEJM 347:10, 2002
HIV Superinfection
 Evidence supports clades from different
geographic areas have combined
 Likely due to superinfection of an individual
harboring a virus of one clade with a second
virus of another clade
McCutchan et al, 1996:N AIDS 10: supp
Robertson et al, 1995: J Mol Evol: 40
SIV Superinfection
 SIV superinfection in monkeys may occur, probably
rare
 Difficult to superinfect a monkey with established
SIV even with
 High infectious dose
 IV administration
 Possible when challenged with second SIV strain
during or soon after initial infection with first strain
 Possible “window of opportunity” for superinfection
Sharpe et al, 1997: J Gen Virol: 78
SIV Superinfection
 Development of virus-specific immunity over
time
 Primary infection: immunity absent or too
immature to effectively prevent infection
 Strengthening of virus-specific immune
responses  superinfection less likely
Sharpe et al, 1997: J Gen Virol: 78
Superinfection
Implications for HC Providers
 Consider the possibility of superinfection
 Counsel patients regarding sexual practices
and safer sex
Summary
 Significant advances in assays to detect HIV
infection
 Alternatives to standard EIA/WB testing may
facilitate improved, ongoing HIV screening
 Detection of acute HIV infection needs to enhanced
 Early intervention in acute HIV infection may have
clinical benefits
 Superinfection needs to be considered
 Risk reduction counseling must be ongoing