Alternatives for acute HIV testing: RNA, p24, heat

dissociated p24

Christopher D. Pilcher, MD

University of North Carolina at Chapel Hill

Center for AIDS Research

AcknowledgementsCIPRASUL-University of Caxias do

Sul (UCS), Porto Alegre, Brazil Ricardo deSouza

Emory UniversityFrances PriddyCarlos del Rio

The UNC Project- Lilongwe, MalawiFrancis E.A. MartinsonPeter N. KazembeDavid ChilongoziClement MapanjePhyllita KumweraSyze GamaDave Namakwa

NC DHHSEvelyn FoustJ. Todd McPhersonRhonda AshbyDel Williams

UNC-Chapel Hill Myron S. CohenPeter A. LeoneIrving F. HoffmanSusan A. FiscusJoAnn KurucLisa HightowMatthew A. Price Joseph J. Eron, JrWilliam C. MillerMelissa KerkauPriya JoshiTrang Q. NguyenKristen HamptonMarc SerrePaul AlabanzaAmy JamesBrant Stalzer

NIMH , NIDDK, HPTN, UNC Fogarty Center, UNC STD CRC, UNC CFAR

• Acute HIV infections (first 2-3 months) are estimated to account for as much as half of all HIV transmission (Wawer JID in press)

• They represent 0-10% of detectable infections presenting for HIV testing, depending on the population

• Recognition of the acutely infected population creates opportunities for highly targeted treatment, prevention and surveillance activities

• Diagnosis requires modification of “standard” treatment algorithms

Testing to Identify Acute HIV

HIV NegativeEstablishedHIV Positive

Ab screen

NAATscreen

+

-+

HIV Indeterminate,Possible Acute HIV

-

1. Ab testing up front—reflexed to NAAT if negative

Ab confirm+ -

HIV NegativeEstablishedHIV Positive

NAAT or Ag and Ab screen+

HIV Indeterminate,Possible Acute HIV

-

2. Simultaneous acute and Ab screening—reflexed to Ab confirmation if positive

Ab confirm+ -

Algorithms must include:

• A screening assay for acute HIV infection – EARLY sensitivity for viral nucleic acid or proteins– High specificity to reduce “false alarms”

• Screening and confirmatory testing for established HIV – LATER sensitivity will allow one to discern more acute

infections from established infections

Testing to Identify Acute HIV

Individual specimens

Pools of 10

The gold standard for acute screening: RNA group testing of Ab- specimens

Pilcher, CD et al. JAMA 2002;288:216-221

Pooling schema

A B C D E

Individual specimens

N=100

10 Pools of 10 F G H I J

A B C D E F G H I J

1 Screening Pool

2-Stage Pooling

Individual specimens

N=100

10 Pools of 10

A B C D E F G H I J

A B C D E F G H I J

A

Individual testing on 10 specimens

10 pools of 10 screened

20 Screening Pools Tested

N=2000

Resolution Testing

HIV Negative

Established HIV Positive

+

+-

HIV Indeterminate,Possible Acute HIV

-

NC DHHS STAT Program:all publicly funded VCT in NC (N~120,000 per yr)

90:10:1 pooling

NucliSensNAAT

Vironostika 1st gen EIA

WB

EstablishedHIV Positive

NAAT

+

+

-

F/U Testing(Ab+NAAT) - +

Acute HIVHIV Negative

-

STAT Confirmatory Testing

Vironostika 1st gen EIA

WB

Specimen Pooling/RNA Testing: NC’s STAT Program

• Of 109,250 previously negative clients tested 11/02-10/03, 23/606 (4%) of HIV infections were acute

• 16/23 (67%) acutes were identified in STD clinic testing sites

• 1 pregnant female was identified

• NAAT PPV 90%; Specificity >99.99%

Pooling and NAAT:Logistical and Cost Issues

• Ideally suited to high throughput, centralized testing

• Most efficient at low prevalences of acute HIV

• Turnaround time: 10-14 d

• Cost reasonable

– $3.63 per HIV test performed

– $17,000 per acute index case identified

Evaluating Alternative Algorithms

• 3 studies conducted in high HIV incidence testing populations

• RNA group testing as the gold standard

• Evaluate different algorithms

EstablishedHIV Positive

NAAT

+

+

-

F/U Testing(Ab+NAAT) - +

Acute HIVHIV Negative

-

STAT Testing

Vironostika 1st gen EIA

WB

EstablishedHIV Positive

+

+

- +

Acute HIVHIV Negative

-

p24 vs. Up24 vs. NAAT:n=1440 STD clients in Lilongwe, Malawi

Determine RTUnigold RT

- or discordantNAATp24

Up24

F/U TestingEIA/WB

Lilongwe, Malawi; N=1,440

• Overall, 575 cases of HIV detected

• 20 (3.5%) were WB-/indeterminate and seroconverted

– 12/16 (75%) detected by standard p24 Ag

– 16/19 (84%) detected by Up24 Ag

– 20/20 (100%) detected by NAAT

Performance of p24 and Up24?Fiscus, et al 12th CROI

Estimate (95% CI)

Sensitivity p24 0.750 (0.476, 0.927)

Sensitivity Up24 0.842 (0.604, 0.966)

Specificity 0.995 (0.988, 0.999)

LR+ p24 161.8 (58.5, 447.8)

LR+ Up24 181.7 (67.0, 492.3)

LR- 0.25 (0.11, 0.59)

p24 and post-test probability: Malawi STD clinic

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

0 0.005 0.01 0.015 0.03 0.035 0.04

Prevalence

Po

st-t

est

Pro

bab

ility

p24

Up24

Negative

0.0226

Expected post-test prob for p24+ (prev .023):

0.79

p24 and post-test probability: Urban US STD Clinic

0

0.1

0.2

0.3

0.4

0.5

0.6

0.7

0.8

0.9

1

0 0.001 0.01 0.015 0.03 0.035 0.04

Prevalence

Po

st-t

est

Pro

bab

ility

p24

Up24

Negative

0.0226

Expected post-test prob for p24+ (prev 0.001):

0.15

+ -

+

-

The 4th generation EIA vs. NAATN=933 VCT clients in Porto Alegre, Brazil

Genscreen Biorad HIV Ag/Ab EIA

++

HIV Negative183 ESTABLISHED HIV

97.3% all HIV+5 ACUTE HIV

2.7 % all HIV+

NAAT

-Biomanguinhos IF

Abbott MEIA

4th generation EIA performance: preliminary observations

• 5/5 (100%) acutes detected consistent with previous observations of p24 sensitivity

• Specificity for acute HIV needs further evaluation

Summary: p24 performance

• Algorithms based on p24-based screening (std p24, Up24, 4th generation EIA) have adequate sensitivity and specificity for acute HIV screening in settings with ~1% or greater prevalence of acute HIV infection (e.g., African STD clinics)

• However, the risk of false positives and costs of individual re-testing all Ab negative specimens may be unacceptable in most US routine testing populations

Laboratory Detection of HIVfrom Pilcher CD, et al. JCI 2004

0 1 2 3 4 5 6 7 8 9 10

Symptoms (~60%)p24 AntigenHIV RNA

HIV Ab Tests

Weeks Since Infection

1st generation (viral lysate) EIAs

Laboratory Detection of HIVfrom Pilcher CD, et al. JCI 2004

0 1 2 3 4 5 6 7 8 9 10

Symptoms (~60%)p24 AntigenHIV RNA

HIV Ab Tests

Weeks Since Infection

3rd generation EIA, MEIA, rapid tests

1st generation (viral lysate) EIAs

+ -

+

-

deSouza, et al unpublished N=933 VCT clients in Porto Alegre, Brazil

Genscreen Biorad HIV Ag/Ab EIA

++

HIV Negative183 ESTABLISHED HIV

97.3% all HIV+5 ACUTE HIV

2.7 % all HIV+

NAAT

-Biomanguinhos IF

Abbott MEIA

68 ESTABLISHED HIV(1 false- rLAV assay)

95.8% all HIV+

NAAT

+

+

- or indeterm.

F/U TestingEIA/WB- +

HIV negative

-

Priddy, et al. 12th CROIn=2,202 VCT and STD clients in Atlanta, GA

Gen Systems HIV ½+O EIA

WB

3 ACUTE HIV

4.2 % all HIV+

555 ESTABLISHED HIV(2 false – on RTs)

96.5% all HIV+

+

20 ACUTE HIV

3.5% all HIV+HIV Negative

Fiscus, et al. 12th CROIn=1440 STD clients in Lilongwe, Malawi

Determine RTUnigold RT

- or discordant (n=22)

+

- +

-NAAT

F/U TestingEIA/WB

Rapid Test Sensitivity

• In a Malawi STD clinic with a high ratio of incidence-to-prevalence, the sensitivity of rapid tests for HIV infection varied with their sensitivity for acute HIV– Determine: detected 559 of 575 (97.2%) – Unigold: detected 554 of 575 (96.3%)

• Performance should be similar in a US STD clinic or MSM testing population

• 6 (30%) of 20 WB -/indeterminate acute HIV infections were Determine+ but Unigold –

• Choice of the rapid test, or series of rapid tests used, can significantly affect the ability to detect early infections

Summary: Ab Screening Choices• Even maximum-sensitivity antibody tests will miss

between 2-5% of detectable HIV infections in high risk testing populations

• If acute screening is to be done, maximum early-sensitivity Ab tests are neither necessary nor desirable

• If acute screening is not to be done, choice of Ab assay can significantly affect HIV detection in testing populations with high proportion of incident cases