1. Testing for Acute HIV Infection in
North Carolina
Myra Brinson, MT(ASCP)
Virology/Serology Unit Manager
North Carolina State Laboratory of Public Health
Ph: 919-807-8835
E-mail: Myra.Brinson@dhhs.nc.gov

2. Background
 A significant portion of all HIV infections in a US
routine testing population are missed by routine HIV
Ab testing.
 HIV-1 viral RNA is present in large copy numbers
during acute infection.
 With multistage pooling, NAATs can efficiently
diagnose acute infection with good positive predictive
value in low prevalence populations.
 It is feasible for laboratories with high testing volume
such as commercial and state public health labs to
perform widespread screening for acute infection.

3. HIV-1 Testing Timeline:
Symptoms
p24 Antigen
HIV RNA
HIV ELISA
0 1 2 3 4 5 6 7 8 9 10
Weeks Since Infection

4. 2001 Pilot Study Design
 All consecutive routine HIV
tests submitted to the NC
State Laboratory of Public
Health over 4 weeks from 110
publicly funded counseling and
Figure 1: Schema for pooling Ab-negative specimens
testing sites (CTS) [n=8505]
Individual Specimens Intermediate Pools Master Pool
 Initial Ab testing - OT
(1:10) (1:90)
Vironostika HIV-1 Viral Lysate
Microelisa (State Lab)
 Manual pooling of Ab NR
samples (State Lab)
 Roche Amplicor HIV-1 Monitor
(UNC) – Standard and US

5. 2001 Pilot Study Results
Figure 2 : Disposition of Specimens
 Acute infection: 5 per 10,000
8505 Total Specimens 12 Previously
 Chronic infection: 44 per Tested HIV+
10,000 152 with Data
Incomplete or 2 HIV+, Status
 Overall specificity: 99.99% Questionable Unknown
8341 Persons at Risk 38 EIA Repeat
 Estimated additional cost per Reactive Sera
37 Newly
Diagnosed
specimen: $2.01 WB+ Chronic
8298 EIA negative & 5 1 WB Negative Infections
 Estimated total testing EIA Unconfirmed Pos. v
costs/additional case 1 False +
diagnosed: $4,109 299 Specimens RNA Test
Insufficient Vol.
4 True Positive
Acute
8005 Ab Neg. Specimens Pooled 5 RNA Positive Infections
Pilcher CD et al, JAMA, Vol. 288/No. 2, July 10, 2002

6. Implementation of NC STAT
 Since 2002, NC has identified AHI using NAAT
through the NC Screening & Tracing Active
Transmission (STAT) program
 Statewide collaboration to detect AHI in publicly
funded testing sites
 240,000 HIV samples/year
 2 to 3 week TAT for test results
 Rapid notification/confirmatory testing
 Rapid tracing/prospective screening of partners

7. The STAT Program: Goals
Identify acute HIV infections in the routine HIV CTS population in NC
 Individual health  Public Health
 Improve prognosis with  Recognized previously
acute treatment? missed infections
 Early entry into care and  Access sexual networks:
prevention services • Identify groups being
 Opportunity to avoid actively infected in a
unwitting transmission to community
partners • Identify partners at high
proximate risk of
infection
• Identify
individuals/groups
actively transmitting in a
community
• Maximize impact of all
interventions to reduce
transmission

8. STAT Testing at NCSLPH
HIV-1/HIV-2 Plus O
EIA Antibody Assay (BioRad)
Vironstika HIV-1 enzyme immunoassay (bioMérieux)
APTIMA HIV-1 RNA
Qualitative Assay (GenProbe)
NucliSENS EasyQ®
HIV-1 assay (bioMérieux)
Procleix HIV-1
assay (GenProbe)
NucliSENS
HIV-1 QL assay
(bioMérieux)

9. All clients presenting at approx. 135 publicly-funded
voluntary testing & counseling (VCT) sites are
tested for HIV RNA using specimen pooling
HIV antibody negative
Screen for antibodies to serum samples are
HIV-1, HIV-2 Plus O pooled for HIV-1 NAAT
using BioRad EIA assay using Hamilton STARlet
with EVOLIS automation

10. Pooling Schema
80 HIV-1/HIV-2 Plus O
Nonreactive Samples
1 B Pool (containing 120 µl x
10 A Pools) = 1200 µl
10 A Pools (containing 20 µl x 8
samples) = 160 µl
X X X X X X X X
X X X X X X X X

11. GenProbe APTIMA HIV-1 NAAT
RNA assay
 FDA-cleared HIV-1
RNA qualitative NAAT
for serum/plasma
 Detects HIV-1
subtypes (Groups
M, N, and O)
 Analytical sensitivity:
30 copies/ml (98.5%)
 Clinical specificity:
99.83%

12. NC HIV Testing Algorithm

13. The STAT System
Disease Intervention
State Laboratory Specialist Team
Laboratory Notification, Interviews,
Identification Confirmatory
Testing, Transportation
UNC Weekly Case- to Clinic
Conference
(Surveillance, Lab, DIS, UNC
Evaluation Teams)
Data collection
UNC Acute HIV Program UNC/Duke
Research Database Collaborative
UNC Specimen Repository Free Urgent clinical
evaluation
-surveillance/research testing
Recruitment to studies

14. North Carolina Voluntary Testing and Counseling Testing
Population 2003-2008
No. HIV Tests‡ Chronic Acute
N (%) N (%) N (%)
Total 891,210 -- 3,766 0.42% 125 0.014%
Sex
Male 279,914 31% 2,631 70% 99 79%
Female 600,399 67% 1,076 29% 26 21%
Median Age (Range) 25 (0-99) 33 (0-84) 26 (16-56)
Age
≤21 288,998 32% 524 14% 35 28%
>21 597,395 67% 3,226 86% 90 72%
Race
Black 395,825 44% 2,578 68% 79 63%
White, Non-Hispanic 284,931 32% 663 18% 34 27%
Hispanic 162,901 18% 327 9% 10 8%
American Indian 9,536 1% 24 1% 1 1%
Missing 9,667 1% 34 1% 1 1%
Sexual Risk Group
Female 600,399 67% 1,076 29% 26 21%
Heterosexual Male 231,575 26% 1,084 29% 22 18%
MSM 26,870 3% 1,277 34% 73 58%
Unknown Male 21,469 2% 270 7% 4 3%
‡From NC CTS -- number of HIV Tests performed, previous positives removed.
Note: Table excludes 2 Acute cases identified from 11/1/2002-12/31/2002 as the number of HIV tests and chronic cases
were not available for this 2 month period

15. North Carolina Voluntary Testing and Counseling Testing
Population 2003-2008
No. HIV Tests‡ Chronic Acute
N (%) N (%) N (%)
Total 891,210 -- 3,766 0.42% 125 0.014%
Sex
Male 279,914 31% 2,631 70% 99 79%
Female 600,399 67% 1,076 29% 26 21%
Median Age (Range) 25 (0-99) 33 (0-84) 26 (16-56)
Age
≤21 288,998 32% 524 14% 35 28%
>21 597,395 67% 3,226 86% 90 72%
Race
Black 395,825 44% 2,578 68% 79 63%
White, Non-Hispanic 284,931 32% 663 18% 34 27%
Hispanic 162,901 18% 327 9% 10 8%
American Indian 9,536 1% 24 1% 1 1%
Missing 9,667 1% 34 1% 1 1%
Sexual Risk Group
Female 600,399 67% 1,076 29% 26 21%
Heterosexual Male 231,575 26% 1,084 29% 22 18%
MSM 26,870 3% 1,277 34% 73 58%
Unknown Male 21,469 2% 270 7% 4 3%
‡From NC CTS -- number of HIV Tests performed, previous positives removed.
Note: Table excludes 2 Acute cases identified from 11/1/2002-12/31/2002 as the number of HIV tests and chronic cases
were not available for this 2 month period

16. North Carolina Voluntary Testing and Counseling Testing
Population 2003-2008
No. HIV Tests‡ Chronic Acute
N (%) N (%) N (%)
Total 891,210 -- 3,766 0.42% 125 0.014%
Sex
Male 279,914 31% 2,631 70% 99 79%
Female 600,399 67% 1,076 29% 26 21%
Median Age (Range) 25 (0-99) 33 (0-84) 26 (16-56)
Age
≤21 288,998 32% 524 14% 35 28%
>21 597,395 67% 3,226 86% 90 72%
Race
Black 395,825 44% 2,578 68% 79 63%
White, Non-Hispanic 284,931 32% 663 18% 34 27%
Hispanic 162,901 18% 327 9% 10 8%
American Indian 9,536 1% 24 1% 1 1%
Missing 9,667 1% 34 1% 1 1%
Sexual Risk Group
Female 600,399 67% 1,076 29% 26 21%
Heterosexual Male 231,575 26% 1,084 29% 22 18%
MSM 26,870 3% 1,277 34% 73 58%
Unknown Male 21,469 2% 270 7% 4 3%
‡From NC CTS -- number of HIV Tests performed, previous positives removed.
Note: Table excludes 2 Acute cases identified from 11/1/2002-12/31/2002 as the number of HIV tests and chronic cases
were not available for this 2 month period

17. North Carolina Voluntary Testing and Counseling Testing
Population 2003-2008
No. HIV Tests‡ Chronic Acute
N (%) N (%) N (%)
Total 891,210 -- 3,766 0.42% 125 0.014%
Sex
Male 279,914 31% 2,631 70% 99 79%
Female 600,399 67% 1,076 29% 26 21%
Median Age (Range) 25 (0-99) 33 (0-84) 26 (16-56)
Age
≤21 288,998 32% 524 14% 35 28%
>21 597,395 67% 3,226 86% 90 72%
Race
Black 395,825 44% 2,578 68% 79 63%
White, Non-Hispanic 284,931 32% 663 18% 34 27%
Hispanic 162,901 18% 327 9% 10 8%
American Indian 9,536 1% 24 1% 1 1%
Missing 9,667 1% 34 1% 1 1%
Sexual Risk Group
Female 600,399 67% 1,076 29% 26 21%
Heterosexual Male 231,575 26% 1,084 29% 22 18%
MSM 26,870 3% 1,277 34% 73 58%
Unknown Male 21,469 2% 270 7% 4 3%
‡From NC CTS -- number of HIV Tests performed, previous positives removed.
Note: Table excludes 2 Acute cases identified from 11/1/2002-12/31/2002 as the number of HIV tests and chronic cases
were not available for this 2 month period

18. North Carolina Voluntary Testing and Counseling Testing
Population 2003-2008
No. HIV Tests‡ Chronic Acute
N (%) N (%) N (%)
Total 891,210 -- 3,766 0.42% 125 0.014%
Sex
Male 279,914 31% 2,631 70% 99 79%
Female 600,399 67% 1,076 29% 26 21%
Median Age (Range) 25 (0-99) 33 (0-84) 26 (16-56)
Age
≤21 288,998 32% 524 14% 35 28%
>21 597,395 67% 3,226 86% 90 72%
Race
Black 395,825 44% 2,578 68% 79 63%
White, Non-Hispanic 284,931 32% 663 18% 34 27%
Hispanic 162,901 18% 327 9% 10 8%
American Indian 9,536 1% 24 1% 1 1%
Missing 9,667 1% 34 1% 1 1%
Sexual Risk Group
Female 600,399 67% 1,076 29% 26 21%
Heterosexual Male 231,575 26% 1,084 29% 22 18%
MSM 26,870 3% 1,277 34% 73 58%
Unknown Male 21,469 2% 270 7% 4 3%
‡From NC CTS -- number of HIV Tests performed, previous positives removed.
Note: Table excludes 2 Acute cases identified from 11/1/2002-12/31/2002 as the number of HIV tests and chronic cases
were not available for this 2 month period

19. North Carolina Voluntary Testing and Counseling Testing
Population 2003-2008
No. HIV Tests‡ Chronic Acute
N (%) N (%) N (%)
Total 891,210 -- 3,766 0.42% 125 0.014%
Sex
Male 279,914 31% 2,631 70% 99 79%
Female 600,399 67% 1,076 29% 26 21%
Median Age (Range) 25 (0-99) 33 (0-84) 26 (16-56)
Age
≤21 288,998 32% 524 14% 35 28%
>21 597,395 67% 3,226 86% 90 72%
Race
Black 395,825 44% 2,578 68% 79 63%
White, Non-Hispanic 284,931 32% 663 18% 34 27%
Hispanic 162,901 18% 327 9% 10 8%
American Indian 9,536 1% 24 1% 1 1%
Missing 9,667 1% 34 1% 1 1%
Sexual Risk Group
Female 600,399 67% 1,076 29% 26 21%
Heterosexual Male 231,575 26% 1,084 29% 22 18%
MSM 26,870 3% 1,277 34% 73 58%
Unknown Male 21,469 2% 270 7% 4 3%
‡From NC CTS -- number of HIV Tests performed, previous positives removed.
Note: Table excludes 2 Acute cases identified from 11/1/2002-12/31/2002 as the number of HIV tests and chronic cases
were not available for this 2 month period

20. North Carolina Voluntary Testing and Counseling Testing Population
2003-2008 (cont.)
No. HIV Tests‡ Chronic Acute
N (%) N (%) N (%)
Site Type
CHC/PHC 17,189 2% 109 3% 3 2%
Drug Treatment 9,944 1% 31 1% 0 0%
Family Planning 156,981 18% 61 2% 0 0%
Field Visit 12,819 1% 316 8% 8 6%
HIV CTS 62,056 7% 872 23% 23 18%
Hosp/PMD 138 0% 3 0% 0 0%
Other 43,996 5% 380 10% 15 12%
Outreach 24,043 3% 153 4% 1 1%
Prenatal/OB 148,996 17% 96 3% 2 2%
Prison/Jail 28,836 3% 231 6% 5 4%
STD 348,896 39% 1,386 37% 54 43%
TB 6,872 1% 14 0% 0 0%
Missing 30,439 3% 114 3% 14 11%
‡From NC CTS -- number of HIV Tests performed, previous positives removed.
Note: Table excludes 2 Acute cases identified from 11/1/2002-12/31/2002 as the number of HIV tests and chronic cases
were not available for this 2 month period

21. North Carolina Voluntary Testing and Counseling Testing Population
2003-2008 (cont.)
No. HIV Tests‡ Chronic Acute
N (%) N (%) N (%)
Site Type
CHC/PHC 17,189 2% 109 3% 3 2%
Drug Treatment 9,944 1% 31 1% 0 0%
Family Planning 156,981 18% 61 2% 0 0%
Field Visit 12,819 1% 316 8% 8 6%
HIV CTS 62,056 7% 872 23% 23 18%
Hosp/PMD 138 0% 3 0% 0 0%
Other 43,996 5% 380 10% 15 12%
Outreach 24,043 3% 153 4% 1 1%
Prenatal/OB 148,996 17% 96 3% 2 2%
Prison/Jail 28,836 3% 231 6% 5 4%
STD 348,896 39% 1,386 37% 54 43%
TB 6,872 1% 14 0% 0 0%
Missing 30,439 3% 114 3% 14 11%
‡From NC CTS -- number of HIV Tests performed, previous positives removed.
Note: Table excludes 2 Acute cases identified from 11/1/2002-12/31/2002 as the number of HIV tests and chronic cases
were not available for this 2 month period

22. Initial HIV Testing Patterns of Acute Patients Strati
by EIA Generation Type
2nd Generation† 3rd Generation‡
(2003-2007) (2008)
N (%) N (%)
EIA(-); NAAT(+) 80 85% 17 57%
EIA(+); WB(Ind.) 12 13% 10 33%
EIA(+); WB(-) 2 2% 3 10%
†2nd Generation EIA: Vironstika HIV-1 (bioMérieux)
‡3rd Generation EIA: HIV-1/HIV-2 Plus O (BioRad)

23. HIV Prevalence in Voluntary Testing and
Counseling Testing Population Stratified by
Calendar Year
No. HIV Tests‡ Chronic Acute
N N % N %
Calendar Year
2003 106,994 581 0.54% 22 0.021%
2004 118,885 552 0.46% 21 0.018%
2005 130,440 590 0.45% 21 0.016%
2006 145,396 645 0.44% 15 0.010%
2007 175,341 670 0.38% 16 0.009%
2008 214,154 728 0.34% 30 0.014%
‡From NC CTS -- number of HIV Tests performed, previous positives removed.

24. Conclusions
 In total, 3.2% of infections identified by the STAT
Program for Antibody-plus-RNA HIV testing
were AHI, which would have been missed using
standard antibody based testing procedures.
 The program’s success at identifying new HIV
infections has led to significant policy changes in
NC, including universal reflex RNA testing for
EIA-negative specimens and mandatory re-
testing of pregnant women in the 3rd trimester.

25. Conclusions
 The yield of AHI cases varied by site type and
type of antibody tests performed. Improved
sensitivity of current EIAs has resulted in a
decreased number of cases presenting as EIA (-)
and HIV-1 RNA (+).
 Decreased proportion of infected individuals in
NC testing population may be due to increased
testing among low risk populations following the
adoption of the CDC opt-out testing in 2007.

26. Conclusions
 Testing for AHI is especially important in high risk
populations such as young MSM of color for whom these
infections would have been missed. The recent increase
in AHI detection may be due to increased testing from
targeted outreach programs, to increased AHI/HIV
education, or to increased high risk behaviors such as
internet partner meeting.
 Although nearly 2/3 of people tested for HIV were
women, a smaller percentage of women are detected
during AHI than men. Plausible explanations include
lower perceived risk among clients and providers, and
greater barriers to care and prevention services among
high-risk women.

27. Thank You!
North Carolina State Laboratory of University of North Carolina –
Public Health Chapel Hill
HIV Serology Laboratory Staff JoAnn Kuruc
Ashley Mayo
Cynthia Guy
Joseph Eron
North Carolina Department of
Health and Human Services RTI International
Lynne Sampson Sandra McCoy
John Barnhart
Evelyn Foust University of California –
Rhonda Ashby San Francisco
Peter Leone Chris Pilcher