- Email: [email protected]
Performance of an alternative laboratory-based algorithm for HIV diagnosis in a high-risk population
Journal of Clinical Virology 52S (2011) S5–S10
Contents lists available at SciVerse ScienceDirect
Journal of Clinical Virology journal homepage: www.elsevier.com/locate/jcv
Performance of an alternative laboratory-based algorithm for HIV diagnosis in a high-risk population Kevin P. Delaney a,∗ , James D. Heffelfinger a , Laura G. Wesolowski a , S. Michele Owen a , William A. Meyer III b , Susan Kennedy a , Apurva Uniyal c , Peter R. Kerndt c , Bernard M. Branson a a
Division of HIV/AIDS Prevention, Centers for Disease Control and Prevention, 1600 Clifton Rd NE, MS E-46, Atlanta, GA 30333, United States Quest Diagnostics, 1901 Sulphur Spring Road, Baltimore, MD 21227, United States c STD Control Program, Los Angeles County Department of Health, Los Angeles, CA, United States b
a r t i c l e Keywords: HIV testing Algorithm Sensitivity Specificity HIV RNA assay
i n f o
a b s t r a c t Background: An immunoassay (IA) followed by Western blot (WB) or immunofluorescence assay has been the primary algorithm used to provide laboratory confirmation of the diagnosis of HIV infection in the US for more than 20 years. Recently, an alternative diagnostic algorithm was proposed to more accurately identify early HIV-1 infection and differentiate between HIV-1 and HIV-2 infection. Objectives: Evaluate a sequential alternative algorithm in which reactive IAs are followed by a rapid HIV test and, if negative, a nucleic acid amplification test (NAAT). Study design: Specimens from high-risk persons were tested with 4 HIV IAs, 6 rapid HIV tests and NAAT (APTIMA® ), which are approved by the United States Food and Drug Administration. IAs were repeated in duplicate if specimen volumes were sufficient. The performance of the alternative algorithm was compared to HIV WB and NAAT. Results: The original study classified 377 specimens as HIV-positive and 3070 as HIV-negative. All 4 IAs correctly identified >99.5% of HIV-positive specimens and, on initial screening, >95.8% of HIV-negative specimens. When repeated, specificity of IAs improved to >99%. Between 6.7% and 12.4% of IA-repeatedly reactive specimens required APTIMA for resolution. The alternative algorithm led to the correct classification of all IA-reactive specimens. Conclusions: Regardless of screening IA and rapid test used, the alternative algorithm correctly classified the infection status of all persons with reactive screening IA results. Few specimens required NAAT for resolution, and the proportion requiring NAAT was lower when repeat IA test results were considered. Published by Elsevier B.V.
1. Background The current algorithm for conducting HIV testing has been in place for more than 20 years. It consists of performing an HIV antibody immunoassay (IA), which, if reactive, is followed by Western blot (WB) or immunofluorescence assay (IFA).1 Third-generation IAs, which detect HIV-1/2 antibodies, and fourth-generation IAs, which detect p24 antigen and HIV1/2 antibodies, have better sensitivity during early infection than WB or IFA, which fail to identify early infection.2–4 In a recently proposed5 alternative to the traditional HIV testing algorithm a reactive or repeatedly reactive IA (depending on manufacturer instructions) is followed by supplemental testing with an antibody test approved by the United States
∗ Corresponding author at: 1600 Clifton Rd., Mailstop E-46, Atlanta, GA 30, United States. E-mail address: [email protected] (K.P. Delaney). 1386-6532/$ – see front matter Published by Elsevier B.V. doi:10.1016/j.jcv.2011.09.013
(US) Food and Drug Administration (FDA). If the supplemental antibody test is negative, a nucleic acid amplification test (NAAT) is performed to resolve this discrepancy. This algorithm, as proposed at the 2010 HIV Diagnostics Conference,5 specifies that the supplemental antibody test should be an HIV-1/HIV-2 differentiation test, but only one such test, the MultispotTM HIV-1/HIV-2 Rapid Test (Bio-Rad Laboratories) is currently approved by the FDA.6 Other rapid HIV tests have similar sensitivity and specificity7 and might be suitable as the supplemental antibody test.
2. Objective This study used specimens collected from a population at high risk for HIV infection to assess the performance of an alternative laboratory-based HIV testing algorithm with antibody IAs, rapid tests, and a NAAT that has been approved for diagnosis of HIV by the FDA.
S6
K.P. Delaney et al. / Journal of Clinical Virology 52S (2011) S5–S10
Fig. 1. Schematic showing the construction of 24 combinations of screening immunoassays and supplemental rapid tests into proposed alternative laboratory-based algorithms for HIV diagnosis.
3. Study design The serum and plasma specimens used in this study were originally collected as part of an evaluation of rapid HIV tests from persons seeking HIV testing at two clinics in Los Angeles, CA. All specimens were tested with 6 rapid tests, namely: the OraQuick Advance® HIV 1/2 Antibody test (OraSure Technologies, Bethlehem, PA), Uni-GoldTM Recombigen® HIV-1 (Trinity Biotech, Bray, Ireland), Reveal® G3 Rapid HIV-1 Antibody Test (MedMira, Inc., Nova Scotia, Canada), Multispot HIV-1/HIV-2 Rapid Test (Bio-Rad Laboratories, Redmond, WA), Clearview® HIV 1/2 STAT-PAK and the Clearview® COMPLETE HIV 1/2 (Inverness Medical Diagnostics, Princeton, NJ). These specimens were originally tested with the Vironostika® HIV-1 Microelisa (bioMerieux, Inc., Durham, NC) by the Los Angeles County Department of Public Health Laboratory between June 2003 and August 2005 as previously described.7 Specimens with repeatedly reactive Vironostika results were tested with the GS HIV-1 Western blot (Bio-Rad Laboratories, Redmond, WA). Remnant serum and plasma specimens were stored frozen at −70 ◦ C until subsequent testing using the four IAs and NAAT was performed during August 2008 to April 2010. Specimens with negative Vironostika results were combined into 16 member pools8 and tested with the APTIMA® HIV-1 RNA qualitative test (Gen-Probe, San Diego, CA). This pooling strategy has been shown to reliably detect HIV-1 RNA at levels as low as 1000 copies/mL.8 Remnant plasma from specimens with positive WB results was also tested in singlet according to the APTIMA package insert. APTIMA and the GS HIV-1/2 plus O (BIORAD, Bio-Rad Laboratories, Redmond, WA) were performed at the Centers for Disease Control and Prevention (CDC) HIV laboratory (Atlanta, GA). The HIVAB HIV-1/HIV-2 rDNA IA (ABBOTT, Abbott Laboratories, Des Plaines, IL) was performed at Abbott laboratories. The Ortho VITROS® anti-HIV-1/2
(Ortho Clinical Diagnostics, Raritan, NJ) and Siemens ADVIA Centaur® EHIV (Siemens Healthcare Diagnostics, Deerfield, IL), were performed at a commercial laboratory. All four laboratorybased IAs were performed according to manufacturer instructions except that repeat testing of all initially reactive specimens was not performed, due to limited specimen volume. Testing was repeated for both BIORAD and ABBOTT tests on specimens with sufficient volume if initial IA results were reactive and WB or APTIMA results were negative. The performance of the proposed alternative algorithm was assessed using the results of screening IAs, rapid tests, and APTIMA to create 24 possible test combinations (Fig. 1). Specimens reactive by the screening IA were classified as HIV infected or HIV uninfected according to the alternative algorithm described by Pandori and Branson.5 Performance of the algorithm was assessed using each rapid test as a supplemental test. Specimens with positive supplemental rapid test results were considered HIV-positive; specimens with negative supplemental rapid test results were classified as HIV-positive or negative based on APTIMA results. Results from the alternative algorithm using different combinations of screening IAs and rapid tests with NAAT were compared to those from the traditional laboratory-based HIV testing algorithm. APTIMApositive specimens with negative or indeterminate WB results were considered to represent acute HIV infection (AHI). When volume permitted, AHI specimens were tested using Roche Amplicor HIV-1 Monitor to quantify RNA copies (Table 1). 4. Results Of the 3447 specimens included in this analysis, 167 collected from persons previously known to be infected with HIV-1 were positive by Vironostika and WB, all four IAs, and all 6 rapid HIV tests; therefore, all 167 would have been correctly classified as
K.P. Delaney et al. / Journal of Clinical Virology 52S (2011) S5–S10
S7
Table 1 Results of the Roche Amplicor HIV-1 Monitor compared with 3rd generation antibody screening tests for 10 specimens identified as acute HIV-1 infections based on Vironostika IA results and pooled RNA screening with the APTIMA HIV-1 Qualitative RNA assay. Specimen ID
Roche Amplicor HIV-1 Monitor results HIV-1 RNA copies/mL
Bio-Rad HIV-1/2 + O
Vitros HIV-1/2
Advia Centaur HIV-1/2/O
Abbott rDNA HIV-1/2
1 2 3 4 5 6 7 8 9 10
Positivea Missingb 631,290 Positivea 482,592 222,929 165,996 5787 2140 294
P P N P P N N N N N
P N N P P N N N N N
P N N P P N N N N N
N N N N P N N N N N
IA, immunoassay. a Quantitative standard too low to calculate (>750,000 copies). b No sample available for testing with the Roche Amplicor HIV-1 RNA assay.
HIV-positive by the proposed algorithm. Remaining analyses are limited to 3280 specimens collected from persons whose HIV status was unknown when they enrolled in the study. Of these specimens, 10 (0.3%) had negative Vironostika but positive APTIMA results and were considered to represent AHI. Nine of these 10 AHI specimens had sufficient volume for testing with the Roche Amplicor HIV-1 Monitor, and all had detectable viral RNA (Table 1). Nine of 10 AHI specimens had negative WB results and one had an indeterminate WB result; the 9 specimens with negative WB results would have been considered HIV-negative by the current algorithm. Two hundred (6.1%) of the remaining specimens had positive Vironostika and WB results and 3070 had negative Vironostika and APTIMA results. Table 2 shows the results of the four screening IAs on this panel of specimens. The performance of the four IAs was similar, although the BIORAD assay detected one more AHI specimen and one additional WB-positive specimen than the other IAs. Overall, 129 specimens with negative Vironostika and APTIMA results had positive initial BIORAD results, however, no more than 25 of these specimens had positive initial results on screening with any of the other three IAs. Of 129 specimens with initial false-positive BIORAD results, only 8 (6.4%) of 125 with sufficient specimen for repeat testing had repeatedly reactive results. For the ABBOTT test, 20 of 25 specimens with initial false-positive results had sufficient specimen for repeat testing; 18 of these remained false-positive when retested. A maximum of 333 (204 true-positive, 129 false-positive) specimens reactive on initial screening with the BIORAD assay would
have required a rapid supplemental test, and 225, 218, and 214 specimens would have required supplemental testing after the ABBOTT, VITROS and ADVIA tests, respectively. The number that would require supplemental testing following the BIORAD assay would be reduced from 333 to 212 if only specimens that were repeatedly reactive were included in the analysis. Table 3 shows the results of the rapid tests performed on these specimens, however, not all specimens had results for all rapid tests. The rapid tests correctly classified over 96% of specimens with truepositive IA results (Table 3). One of four AHI specimens detected by the screening IAs had reactive Unigold and Multispot results. Depending on the test combination, between 2 and 7 HIV-infected specimens had reactive IA and negative rapid test results, and all of these specimens were correctly classified as HIV-positive by subsequent APTIMA testing. The only false-negative algorithm results occurred among specimens with negative screening IA results (Table 2). Only two specimens that had false-positive initial IA results had a concordant false-positive rapid test result (both specimens were false-positive on BIORAD and Multispot). When the BIORAD assay was repeated, these two specimens were negative. The positive predictive value (PPV) for all combinations of IAs and rapid tests was 100% (95% confidence interval [CI]: 97.7–100) except for the BIORAD-Multispot combination, for which the PPV, based on initial IA results only, was 99.0% (95% CI: 96.4–99.9). Combining the total number of specimens that had either false- or true-negative rapid tests results, between 7% (14 of 206 for ADVIA-MULTISPOT) and 14% (28 of 203 for ABBOTT-UNIGOLD) of specimens initially
Table 2 Proportions of specimens (N = 3280) collected from high-risk persons seeking HIV testing correctly classified by four HIV immunoassays (IA); Los Angeles, CA June 2005–June 2008. Algorithm of EIA/WB and NAAT
IA and WB positive specimensa IA and APTIMA negative specimensb Acute infection specimensc
Screening antibody immunoassay
N = 200 N = 3070 N = 10
Vitros N (%) correct
Advia N (%) correct
Bio-Rad N (%) correct
199 (99.5) 3054 (99.5) 3 (30.0)
199 (99.5) 3058 (99.6) 3 (30.0)
200 (100) 2941d (95.8) 4(40.0)
Abbott N (%) correct 199 (99.5) 3045e (99.2) 1 (10.0)
NAAT, nucleic acid amplification testing; IA, immunoassay; WB, Western blot. a Specimens with repeatedly reactive Vironostika HIV-1 Immunoassay (Vironostika) results confirmed as HIV-1 positive by the GS HIV-1 Western Blot. Proportion correct corresponds to screening immunoassay sensitivity. b Specimens with negative Vironostika results that also tested negative when pooled in 16-member pools and tested by APTIMA in 16-member pools as described in Ref. [8]. Proportion correct corresponds to the screening immunoassay specificity when run in singlet. c Specimens with negative Vironostika results that tested positive by APTIMA when pooled in 16-member pools. Pools were deconstructed so that all 10 of these specimens had individual positive APTIMA results. d 125 of 129 IA false-positive specimens had sufficient volume for repeat testing. Based on repeatedly reactive IA results, 3058 of 3066 specimens (99.7%) were correctly classified as HIV-negative. e 20 of 25 IA false-positive specimens had sufficient volume for repeat testing. Based on repeatedly reactive IA results 3047 of 3065 specimens (99.4%) were correctly classified as HIV-negative.
N, number; IA, immunoassay. a Specimens positive by the Vironostika IA and Western blot or APTIMA and reactive by the screening immunoassay. b Specimens negative by the Vironostika IA and APTIMA but reactive by the screening immunoassay. c This is the maximum number of specimens with reactive IA results but a missing rapid test result. The number of missing rapid test results for any column can be calculated as column total minus the N with correct and incorrect rapid test results (N/%) for that column. d Specimens correctly categorized by the supplemental rapid HIV test; true positive specimens had positive rapid test results and false-positive specimens had negative rapid test results.
21 (100) 23 (100) 25 (100) 25 (100) 25 (100) 23 (100) 161 (98.8) 178 (98.9) 195 (99.0) 192 (99.0) 196 (98.5) 175 (97.2) 117 (100) 121 (100) 129 (100) 123 (98.4) 129 (100) 121 (100)
N rapid test correct (%)
161 (97.0) 178 (96.7) 195 (97.0) 194 (98.0) 196 (96.6) 177 (96.2) 11 (100) 10 (100) 12 (100) 10 (100) 12 (100) 10 (100)
N rapid test correct (%)
161 (97.6) 178 (97.8) 195 (98.0) 192 (98.0) 196 (97.5) 175 (96.2) 13 (100) 14 (100) 16 (100) 14 (100) 16 (100) 14 (100) 161 (97.6) 178 (97.8) 195 (98.0) 192 (98.0) 196 (97.5) 175 (96.2) 39, 12 20, 8 3, 0 6, 4 1, 0 20, 8 Clearview Complete Clearview Stat-Pak OraQuick Advance Multispot HIV-1/2 Reveal G3 HIV-1 Unigold Recombigen HIV-1
N rapid test correctd (%)
True Positive N = 200 True Positive Na = 202
False Positive Nb = 16
True Positive N = 202
False Positive N = 12
True Positive N = 204
False Positive N = 129
Abbott Bio-Rad Advia Maximum IA True Positive, Vitros IA False Positive Missing Rapid testc Rapid test
Table 3 Performance of rapid tests as supplemental tests following reactive 3rd generation screening immunoassays.
N rapid test correct (%)
K.P. Delaney et al. / Journal of Clinical Virology 52S (2011) S5–S10
False Positive N = 25
S8
reactive on the ABBOTT, VITROS or ADVIA would have required testing with APTIMA. If the ABBOTT test was repeated in duplicate, the proportion requiring APTIMA would have been reduced to 12.8%. For BIORAD, if the initial test result was considered, between 39% (BIORAD-MULTISPOT) and 44% (BIORAD-CLEARVIEW) of initially reactive specimens would have required APTIMA testing. However, if the BIORAD test was repeated as specified in the current FDA-approved package insert, this number would decrease to no more than 15 (7.1%) of 212 specimens, similar to the other screening IAs.
5. Discussion In this study evaluating the proposed alternative algorithm5 among specimens from persons at high risk for HIV infection, all third generation screening IAs had similar sensitivity and specificity, and rapid supplemental tests distinguished the majority of specimens with positive and negative antibody results. There were few instances of false-negative results with the proposed new algorithm, and these were primarily among specimens with AHI. All such specimens had negative results on the initial screening assay. Fourth generation screening assays may have been able to identify a greater proportion of these acute infections due to their ability to detect p24 antigen during the earliest stage of infection, although some (30–40%) AHIs were detected by third generation IAs. There were two instances where the combination of BIORAD followed by Multispot produced false-positive algorithm results. However, if the BIORAD test was repeated, as specified in the package insert, these specimens would have been classified as HIVnegative. The specimens used in this study were prospectively collected from a population with high prevalence (∼4%) of undiagnosed HIV infection7 and a 0.3% yield of AHIs.4 HIV incidence in the United States has been shown to be increasing in some risk groups9 and is highest among men who have sex with men, the risk group from which the majority of specimens in this study were obtained.7 In settings where high-risk persons are tested frequently,3 having a highly sensitive screening test capable of detecting early infection, such as a third or fourth generation IA, is of paramount importance so infected persons do not receive the incorrect information that they are uninfected. Newly developed assays10 are more sensitive for detecting early infection; therefore the number of specimens collected in settings with high HIV incidence that will be negative by WB or IFA and require NAAT testing is expected to increase. The proposed algorithm, which utilizes a rapid supplemental test which is less sensitive for early infection than the screening test,2 will identify more persons in the early stage of infection than would be possible if the screening IA and rapid test had similar seroconversion sensitivities.11 The algorithm would thereby allow for more accurate and rapid distinction of persons with early infection, who are likely to be at high risk for HIV transmission, from persons with undiagnosed infection with lower transmission risk.12,13 In this study, all the screening IAs had similar performance, and therefore other characteristics that we did not evaluate, such as convenience, time-to-results and cost may be important in determining which test methods will be used in different HIV testing settings. For example, both the VITROS and ADVIA assays are run on automated analyzers that allow for loading of individual specimens, are commonly used in healthcare settings to perform a variety of clinical chemistry and serology tests, and provide results in less than an hour. However, use of these analyzers may be cost-prohibitive in some settings due to the costs associated with running appropriate controls for small patient specimen batch sizes. Even after allowing for repeat testing of screening IAs as
K.P. Delaney et al. / Journal of Clinical Virology 52S (2011) S5–S10
specified by manufacturers, using rapid HIV tests after obtaining reactive IA results may expedite the algorithm used to conduct HIV testing and allow more timely confirmation of HIV infection in healthcare settings. Currently, many laboratories that perform HIV screening tests send repeatedly reactive specimens out to reference laboratories for WB testing, increasing the turnaround time for a definitive test result. The proposed algorithm would allow on-site supplemental testing and may thereby obviate some of the challenges to implementation of routine testing for HIV in healthcare setting as recommended by CDC.14 The algorithm proposed at the 2010 HIV Diagnostics Conference5 suggested using an HIV-1/HIV-2 differentiation assay after a reactive HIV-1/HIV-2 screening IA, and others have shown this to be an effective strategy.15–17 In this study we compared the performance of 6 FDA-approved rapid tests for this role. Our finding that all the rapid tests performed similarly as supplemental tests in the algorithm is not unexpected.7 Still, because of the clinical differences in the progression and treatment of HIV-2 disease18 and the lack of an FDA-approved HIV-2 supplemental assay,15 use of an HIV-1/HIV-2 differentiation assay is likely to be the preferred approach. This study has several limitations. Insufficient specimen volumes prevented us from testing all specimens with all test methods. Of particular importance was the inability to subject all specimens with initially reactive IA results to repeat testing with IAs. Repeat testing of the ABBOTT assay had little effect on the number of specimens requiring NAAT. However, repeat testing of the BIORAD assay dramatically reduced the number of specimens with initial falsepositive results that would require a rapid test and NAAT. This is an important finding because it suggests that repeating IAs according to package insert instructions may be more cost-effective than subjecting these additional specimens to rapid testing and NAAT. In this study, we were unable to repeat testing on specimens with initially reactive ADVIA results, in accordance with package insert instructions, and therefore may have underestimated the performance of this test in the algorithm. In addition, the differences in total numbers of specimens tested using different rapid tests interfered with the ability to directly compare the number of NAAT tests required. Another limitation of this study is that none of the specimens collected were from HIV-2 infected individuals and so we could not evaluate how well the proposed algorithm performed at identifying HIV-2 infection. In summary, we found that, using specimens collected from persons at high risk for HIV infection, the proposed laboratory algorithm was able to identify nearly all true infections, including some acute infections and, when tests were performed according to their package insert instructions, produced no false-positive results. This alternative algorithm has the potential to identify both prevalent and acute infection with improved efficiency and fewer false-negative results than the current HIV testing algorithm used in the United States. Funding This work was funded by the Centers for Disease Control and Prevention. Competing interests None declared. Ethical approval The original study under which specimens were obtained from clients seeking HIV testing was reviewed and approved by both CDC
S9
and local institutional review boards. Study participants consented to storage and future testing of stored deidentified specimens, and this testing was subsequently conducted on review by the CDC after being categorized as research not involving identifiable human subjects. Disclaimer The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention or the U.S. Department of Health and Human Services or Quest Diagnostics, Inc. The use of trade names and commercial sources is for identification only and does not imply endorsement by the U.S. Department of Health and Human Services. Acknowledgements The authors would like to acknowledge study staff, especially Lashawnda Royal and Staeci Morita, as well as all the study participants who provided specimens for this study. We would like to acknowledge IA testing by Abbott Diagnostics, as well as the provision of APTIMA testing supplies by GenProbe. We would also like to thank Dollene Hemmerlein, Suzette Bartley, and the staff from the CDC serum bank for receiving, aliquoting and managing storage and shipment of study specimens. References 1. Centers for Disease Control and Prevention. Interpretation and use of the Western blot assay for serodiagnosis of human immunodeficiency virus type 1 infections. MMWR Recomm Rep 1989;38:S-7. 2. Owen SM, Yang C, Spira T, Ou Y, Pau CP, Parekh BS, et al. Alternative algorithms for human immunodeficiency virus infection diagnosis using tests that are licensed in the United States. J Clin Microbiol 2008;46(5): 1588–95. 3. Patel P, MacKellar D, Simmons P, Uniyal A, Gallager K, Bennett B, et al. Detecting acute human immunodeficiency virus infection using three different screening immunoassays and nucleic acid amplification testing for human immunodeficiency virus RNA 2006–2008. Arch Intern Med 2010;170(1): 66–74. 4. Stekler JD, Swenson PD, Coombs RW, Dragavon J, Thomas KK, Brennan CA, et al. HIV testing in a high-incidence population: is antibody testing alone good enough? Clin Infect Dis 2009;49:444–53. 5. Pandori MW, Branson BM. 2010 HIV diagnostics conference. Expert Rev Anti Infect Ther 2010;8(6):631–3. 6. CDC. FDA-approved rapid HIV antibody screening tests. Available from: [accessed http://www.cdc.gov/hiv/topics/testing/rapid/rt-comparison.htm 31.03.11]. 7. Delaney KP, Branson BM, Uniyal A, Phillips S, Candal D, Owen SM, et al. Evaluation of the performance characteristics of 6 rapid HIV antibody tests. Clin Infect Dis 2011;52(2):257–63. 8. Ethridge SF, Hart C, Hanson DL, Parker MM, Bennett B, et al. Performance of the APTIMA HIV-1 RNA qualitative assay with 16- and 32-member specimen pools. J Clin Microbiol 2010;48(9):3343–5. 9. Hall HI, Song R, Rhodes P, Prejean J, An Q, Lee LM, et al. Estimation of HIV incidence in the United States. JAMA 2008;300(5):520–9. 10. Chavez P, Wesolowski L, Patel P, Delaney K, Owen SM. Evaluation of the performance of the Abbott ARCHITECT HIV Ag/Ab Combo Assay. J Clinical Virol 2011;52(Suppl 1):S51–5. 11. Fiscus SA, Pilcher CD, Miller WC, Powers KA, Hoffman IF, Price M, et al. Rapid, real-time detection of acute HIV infection in patients in Africa. J Infect Dis 2007;195(3):416–24. 12. Wawer MJ, Gray RH, Sewankambo NK, Serwadda D, Li X, Laeyendecker O, et al. Rates of HIV-1 transmission per coital act, by stage of HIV-1 infection, in Rakai, Uganda. J Infect Dis 2005;191(May (9)):1403–9. 13. Hollingsworth TD, Anderson RM, Fraser C. HIV-1 transmission, by stage of infection. J Infect Dis 2008;198(September (5)):687–93. 14. CDC. Revised recommendations for HIV testing of adults, adolescents, and pregnant women in health-care settings. MMWR Recomm Rep 2006;55(RR14):1–17. 15. Nasrullah M, Ethridge SF, Delaney KP, Wesolowski LG, Granade TC, Schwendemann J, et al. Comparison of alternative interpretive criteria for the HIV-1
S10
K.P. Delaney et al. / Journal of Clinical Virology 52S (2011) S5–S10
Western blot and results of the Multispot HIV-1/HIV-2 Rapid Test for classifying HIV-1 and HIV-2 infections. J Clin Virol 2011;52(Suppl 1):S23–7. 16. Steyer LM, Sullivan TJ, Parker MM. Evaluation of an alternative supplemental testing strategy for HIV diagnosis by retrospective analysis of clinical HIV testing data. J Clin Virol 2011;52(Suppl 1):S35– 40.
17. Torian LV, Forgione LA, Punsalang AE, Pirillo RE, Oleszko WR. Comparison of Multispot EIA with Western blot for confirmatory serodiagnosis of HIV. J Clin Virol 2011;52(Suppl 1):S41–4. 18. Ntemgwa ML, d’Aquin Toni T, Brenner BG, Camacho RJ, Wainberg MA. Antiretroviral drug resistance in human immunodeficiency virus type 2. Antimicrob Agents Chemother 2009;53(9):3611–9.
Contents lists available at SciVerse ScienceDirect
Journal of Clinical Virology journal homepage: www.elsevier.com/locate/jcv
Performance of an alternative laboratory-based algorithm for HIV diagnosis in a high-risk population Kevin P. Delaney a,∗ , James D. Heffelfinger a , Laura G. Wesolowski a , S. Michele Owen a , William A. Meyer III b , Susan Kennedy a , Apurva Uniyal c , Peter R. Kerndt c , Bernard M. Branson a a
Division of HIV/AIDS Prevention, Centers for Disease Control and Prevention, 1600 Clifton Rd NE, MS E-46, Atlanta, GA 30333, United States Quest Diagnostics, 1901 Sulphur Spring Road, Baltimore, MD 21227, United States c STD Control Program, Los Angeles County Department of Health, Los Angeles, CA, United States b
a r t i c l e Keywords: HIV testing Algorithm Sensitivity Specificity HIV RNA assay
i n f o
a b s t r a c t Background: An immunoassay (IA) followed by Western blot (WB) or immunofluorescence assay has been the primary algorithm used to provide laboratory confirmation of the diagnosis of HIV infection in the US for more than 20 years. Recently, an alternative diagnostic algorithm was proposed to more accurately identify early HIV-1 infection and differentiate between HIV-1 and HIV-2 infection. Objectives: Evaluate a sequential alternative algorithm in which reactive IAs are followed by a rapid HIV test and, if negative, a nucleic acid amplification test (NAAT). Study design: Specimens from high-risk persons were tested with 4 HIV IAs, 6 rapid HIV tests and NAAT (APTIMA® ), which are approved by the United States Food and Drug Administration. IAs were repeated in duplicate if specimen volumes were sufficient. The performance of the alternative algorithm was compared to HIV WB and NAAT. Results: The original study classified 377 specimens as HIV-positive and 3070 as HIV-negative. All 4 IAs correctly identified >99.5% of HIV-positive specimens and, on initial screening, >95.8% of HIV-negative specimens. When repeated, specificity of IAs improved to >99%. Between 6.7% and 12.4% of IA-repeatedly reactive specimens required APTIMA for resolution. The alternative algorithm led to the correct classification of all IA-reactive specimens. Conclusions: Regardless of screening IA and rapid test used, the alternative algorithm correctly classified the infection status of all persons with reactive screening IA results. Few specimens required NAAT for resolution, and the proportion requiring NAAT was lower when repeat IA test results were considered. Published by Elsevier B.V.
1. Background The current algorithm for conducting HIV testing has been in place for more than 20 years. It consists of performing an HIV antibody immunoassay (IA), which, if reactive, is followed by Western blot (WB) or immunofluorescence assay (IFA).1 Third-generation IAs, which detect HIV-1/2 antibodies, and fourth-generation IAs, which detect p24 antigen and HIV1/2 antibodies, have better sensitivity during early infection than WB or IFA, which fail to identify early infection.2–4 In a recently proposed5 alternative to the traditional HIV testing algorithm a reactive or repeatedly reactive IA (depending on manufacturer instructions) is followed by supplemental testing with an antibody test approved by the United States
∗ Corresponding author at: 1600 Clifton Rd., Mailstop E-46, Atlanta, GA 30, United States. E-mail address: [email protected] (K.P. Delaney). 1386-6532/$ – see front matter Published by Elsevier B.V. doi:10.1016/j.jcv.2011.09.013
(US) Food and Drug Administration (FDA). If the supplemental antibody test is negative, a nucleic acid amplification test (NAAT) is performed to resolve this discrepancy. This algorithm, as proposed at the 2010 HIV Diagnostics Conference,5 specifies that the supplemental antibody test should be an HIV-1/HIV-2 differentiation test, but only one such test, the MultispotTM HIV-1/HIV-2 Rapid Test (Bio-Rad Laboratories) is currently approved by the FDA.6 Other rapid HIV tests have similar sensitivity and specificity7 and might be suitable as the supplemental antibody test.
2. Objective This study used specimens collected from a population at high risk for HIV infection to assess the performance of an alternative laboratory-based HIV testing algorithm with antibody IAs, rapid tests, and a NAAT that has been approved for diagnosis of HIV by the FDA.
S6
K.P. Delaney et al. / Journal of Clinical Virology 52S (2011) S5–S10
Fig. 1. Schematic showing the construction of 24 combinations of screening immunoassays and supplemental rapid tests into proposed alternative laboratory-based algorithms for HIV diagnosis.
3. Study design The serum and plasma specimens used in this study were originally collected as part of an evaluation of rapid HIV tests from persons seeking HIV testing at two clinics in Los Angeles, CA. All specimens were tested with 6 rapid tests, namely: the OraQuick Advance® HIV 1/2 Antibody test (OraSure Technologies, Bethlehem, PA), Uni-GoldTM Recombigen® HIV-1 (Trinity Biotech, Bray, Ireland), Reveal® G3 Rapid HIV-1 Antibody Test (MedMira, Inc., Nova Scotia, Canada), Multispot HIV-1/HIV-2 Rapid Test (Bio-Rad Laboratories, Redmond, WA), Clearview® HIV 1/2 STAT-PAK and the Clearview® COMPLETE HIV 1/2 (Inverness Medical Diagnostics, Princeton, NJ). These specimens were originally tested with the Vironostika® HIV-1 Microelisa (bioMerieux, Inc., Durham, NC) by the Los Angeles County Department of Public Health Laboratory between June 2003 and August 2005 as previously described.7 Specimens with repeatedly reactive Vironostika results were tested with the GS HIV-1 Western blot (Bio-Rad Laboratories, Redmond, WA). Remnant serum and plasma specimens were stored frozen at −70 ◦ C until subsequent testing using the four IAs and NAAT was performed during August 2008 to April 2010. Specimens with negative Vironostika results were combined into 16 member pools8 and tested with the APTIMA® HIV-1 RNA qualitative test (Gen-Probe, San Diego, CA). This pooling strategy has been shown to reliably detect HIV-1 RNA at levels as low as 1000 copies/mL.8 Remnant plasma from specimens with positive WB results was also tested in singlet according to the APTIMA package insert. APTIMA and the GS HIV-1/2 plus O (BIORAD, Bio-Rad Laboratories, Redmond, WA) were performed at the Centers for Disease Control and Prevention (CDC) HIV laboratory (Atlanta, GA). The HIVAB HIV-1/HIV-2 rDNA IA (ABBOTT, Abbott Laboratories, Des Plaines, IL) was performed at Abbott laboratories. The Ortho VITROS® anti-HIV-1/2
(Ortho Clinical Diagnostics, Raritan, NJ) and Siemens ADVIA Centaur® EHIV (Siemens Healthcare Diagnostics, Deerfield, IL), were performed at a commercial laboratory. All four laboratorybased IAs were performed according to manufacturer instructions except that repeat testing of all initially reactive specimens was not performed, due to limited specimen volume. Testing was repeated for both BIORAD and ABBOTT tests on specimens with sufficient volume if initial IA results were reactive and WB or APTIMA results were negative. The performance of the proposed alternative algorithm was assessed using the results of screening IAs, rapid tests, and APTIMA to create 24 possible test combinations (Fig. 1). Specimens reactive by the screening IA were classified as HIV infected or HIV uninfected according to the alternative algorithm described by Pandori and Branson.5 Performance of the algorithm was assessed using each rapid test as a supplemental test. Specimens with positive supplemental rapid test results were considered HIV-positive; specimens with negative supplemental rapid test results were classified as HIV-positive or negative based on APTIMA results. Results from the alternative algorithm using different combinations of screening IAs and rapid tests with NAAT were compared to those from the traditional laboratory-based HIV testing algorithm. APTIMApositive specimens with negative or indeterminate WB results were considered to represent acute HIV infection (AHI). When volume permitted, AHI specimens were tested using Roche Amplicor HIV-1 Monitor to quantify RNA copies (Table 1). 4. Results Of the 3447 specimens included in this analysis, 167 collected from persons previously known to be infected with HIV-1 were positive by Vironostika and WB, all four IAs, and all 6 rapid HIV tests; therefore, all 167 would have been correctly classified as
K.P. Delaney et al. / Journal of Clinical Virology 52S (2011) S5–S10
S7
Table 1 Results of the Roche Amplicor HIV-1 Monitor compared with 3rd generation antibody screening tests for 10 specimens identified as acute HIV-1 infections based on Vironostika IA results and pooled RNA screening with the APTIMA HIV-1 Qualitative RNA assay. Specimen ID
Roche Amplicor HIV-1 Monitor results HIV-1 RNA copies/mL
Bio-Rad HIV-1/2 + O
Vitros HIV-1/2
Advia Centaur HIV-1/2/O
Abbott rDNA HIV-1/2
1 2 3 4 5 6 7 8 9 10
Positivea Missingb 631,290 Positivea 482,592 222,929 165,996 5787 2140 294
P P N P P N N N N N
P N N P P N N N N N
P N N P P N N N N N
N N N N P N N N N N
IA, immunoassay. a Quantitative standard too low to calculate (>750,000 copies). b No sample available for testing with the Roche Amplicor HIV-1 RNA assay.
HIV-positive by the proposed algorithm. Remaining analyses are limited to 3280 specimens collected from persons whose HIV status was unknown when they enrolled in the study. Of these specimens, 10 (0.3%) had negative Vironostika but positive APTIMA results and were considered to represent AHI. Nine of these 10 AHI specimens had sufficient volume for testing with the Roche Amplicor HIV-1 Monitor, and all had detectable viral RNA (Table 1). Nine of 10 AHI specimens had negative WB results and one had an indeterminate WB result; the 9 specimens with negative WB results would have been considered HIV-negative by the current algorithm. Two hundred (6.1%) of the remaining specimens had positive Vironostika and WB results and 3070 had negative Vironostika and APTIMA results. Table 2 shows the results of the four screening IAs on this panel of specimens. The performance of the four IAs was similar, although the BIORAD assay detected one more AHI specimen and one additional WB-positive specimen than the other IAs. Overall, 129 specimens with negative Vironostika and APTIMA results had positive initial BIORAD results, however, no more than 25 of these specimens had positive initial results on screening with any of the other three IAs. Of 129 specimens with initial false-positive BIORAD results, only 8 (6.4%) of 125 with sufficient specimen for repeat testing had repeatedly reactive results. For the ABBOTT test, 20 of 25 specimens with initial false-positive results had sufficient specimen for repeat testing; 18 of these remained false-positive when retested. A maximum of 333 (204 true-positive, 129 false-positive) specimens reactive on initial screening with the BIORAD assay would
have required a rapid supplemental test, and 225, 218, and 214 specimens would have required supplemental testing after the ABBOTT, VITROS and ADVIA tests, respectively. The number that would require supplemental testing following the BIORAD assay would be reduced from 333 to 212 if only specimens that were repeatedly reactive were included in the analysis. Table 3 shows the results of the rapid tests performed on these specimens, however, not all specimens had results for all rapid tests. The rapid tests correctly classified over 96% of specimens with truepositive IA results (Table 3). One of four AHI specimens detected by the screening IAs had reactive Unigold and Multispot results. Depending on the test combination, between 2 and 7 HIV-infected specimens had reactive IA and negative rapid test results, and all of these specimens were correctly classified as HIV-positive by subsequent APTIMA testing. The only false-negative algorithm results occurred among specimens with negative screening IA results (Table 2). Only two specimens that had false-positive initial IA results had a concordant false-positive rapid test result (both specimens were false-positive on BIORAD and Multispot). When the BIORAD assay was repeated, these two specimens were negative. The positive predictive value (PPV) for all combinations of IAs and rapid tests was 100% (95% confidence interval [CI]: 97.7–100) except for the BIORAD-Multispot combination, for which the PPV, based on initial IA results only, was 99.0% (95% CI: 96.4–99.9). Combining the total number of specimens that had either false- or true-negative rapid tests results, between 7% (14 of 206 for ADVIA-MULTISPOT) and 14% (28 of 203 for ABBOTT-UNIGOLD) of specimens initially
Table 2 Proportions of specimens (N = 3280) collected from high-risk persons seeking HIV testing correctly classified by four HIV immunoassays (IA); Los Angeles, CA June 2005–June 2008. Algorithm of EIA/WB and NAAT
IA and WB positive specimensa IA and APTIMA negative specimensb Acute infection specimensc
Screening antibody immunoassay
N = 200 N = 3070 N = 10
Vitros N (%) correct
Advia N (%) correct
Bio-Rad N (%) correct
199 (99.5) 3054 (99.5) 3 (30.0)
199 (99.5) 3058 (99.6) 3 (30.0)
200 (100) 2941d (95.8) 4(40.0)
Abbott N (%) correct 199 (99.5) 3045e (99.2) 1 (10.0)
NAAT, nucleic acid amplification testing; IA, immunoassay; WB, Western blot. a Specimens with repeatedly reactive Vironostika HIV-1 Immunoassay (Vironostika) results confirmed as HIV-1 positive by the GS HIV-1 Western Blot. Proportion correct corresponds to screening immunoassay sensitivity. b Specimens with negative Vironostika results that also tested negative when pooled in 16-member pools and tested by APTIMA in 16-member pools as described in Ref. [8]. Proportion correct corresponds to the screening immunoassay specificity when run in singlet. c Specimens with negative Vironostika results that tested positive by APTIMA when pooled in 16-member pools. Pools were deconstructed so that all 10 of these specimens had individual positive APTIMA results. d 125 of 129 IA false-positive specimens had sufficient volume for repeat testing. Based on repeatedly reactive IA results, 3058 of 3066 specimens (99.7%) were correctly classified as HIV-negative. e 20 of 25 IA false-positive specimens had sufficient volume for repeat testing. Based on repeatedly reactive IA results 3047 of 3065 specimens (99.4%) were correctly classified as HIV-negative.
N, number; IA, immunoassay. a Specimens positive by the Vironostika IA and Western blot or APTIMA and reactive by the screening immunoassay. b Specimens negative by the Vironostika IA and APTIMA but reactive by the screening immunoassay. c This is the maximum number of specimens with reactive IA results but a missing rapid test result. The number of missing rapid test results for any column can be calculated as column total minus the N with correct and incorrect rapid test results (N/%) for that column. d Specimens correctly categorized by the supplemental rapid HIV test; true positive specimens had positive rapid test results and false-positive specimens had negative rapid test results.
21 (100) 23 (100) 25 (100) 25 (100) 25 (100) 23 (100) 161 (98.8) 178 (98.9) 195 (99.0) 192 (99.0) 196 (98.5) 175 (97.2) 117 (100) 121 (100) 129 (100) 123 (98.4) 129 (100) 121 (100)
N rapid test correct (%)
161 (97.0) 178 (96.7) 195 (97.0) 194 (98.0) 196 (96.6) 177 (96.2) 11 (100) 10 (100) 12 (100) 10 (100) 12 (100) 10 (100)
N rapid test correct (%)
161 (97.6) 178 (97.8) 195 (98.0) 192 (98.0) 196 (97.5) 175 (96.2) 13 (100) 14 (100) 16 (100) 14 (100) 16 (100) 14 (100) 161 (97.6) 178 (97.8) 195 (98.0) 192 (98.0) 196 (97.5) 175 (96.2) 39, 12 20, 8 3, 0 6, 4 1, 0 20, 8 Clearview Complete Clearview Stat-Pak OraQuick Advance Multispot HIV-1/2 Reveal G3 HIV-1 Unigold Recombigen HIV-1
N rapid test correctd (%)
True Positive N = 200 True Positive Na = 202
False Positive Nb = 16
True Positive N = 202
False Positive N = 12
True Positive N = 204
False Positive N = 129
Abbott Bio-Rad Advia Maximum IA True Positive, Vitros IA False Positive Missing Rapid testc Rapid test
Table 3 Performance of rapid tests as supplemental tests following reactive 3rd generation screening immunoassays.
N rapid test correct (%)
K.P. Delaney et al. / Journal of Clinical Virology 52S (2011) S5–S10
False Positive N = 25
S8
reactive on the ABBOTT, VITROS or ADVIA would have required testing with APTIMA. If the ABBOTT test was repeated in duplicate, the proportion requiring APTIMA would have been reduced to 12.8%. For BIORAD, if the initial test result was considered, between 39% (BIORAD-MULTISPOT) and 44% (BIORAD-CLEARVIEW) of initially reactive specimens would have required APTIMA testing. However, if the BIORAD test was repeated as specified in the current FDA-approved package insert, this number would decrease to no more than 15 (7.1%) of 212 specimens, similar to the other screening IAs.
5. Discussion In this study evaluating the proposed alternative algorithm5 among specimens from persons at high risk for HIV infection, all third generation screening IAs had similar sensitivity and specificity, and rapid supplemental tests distinguished the majority of specimens with positive and negative antibody results. There were few instances of false-negative results with the proposed new algorithm, and these were primarily among specimens with AHI. All such specimens had negative results on the initial screening assay. Fourth generation screening assays may have been able to identify a greater proportion of these acute infections due to their ability to detect p24 antigen during the earliest stage of infection, although some (30–40%) AHIs were detected by third generation IAs. There were two instances where the combination of BIORAD followed by Multispot produced false-positive algorithm results. However, if the BIORAD test was repeated, as specified in the package insert, these specimens would have been classified as HIVnegative. The specimens used in this study were prospectively collected from a population with high prevalence (∼4%) of undiagnosed HIV infection7 and a 0.3% yield of AHIs.4 HIV incidence in the United States has been shown to be increasing in some risk groups9 and is highest among men who have sex with men, the risk group from which the majority of specimens in this study were obtained.7 In settings where high-risk persons are tested frequently,3 having a highly sensitive screening test capable of detecting early infection, such as a third or fourth generation IA, is of paramount importance so infected persons do not receive the incorrect information that they are uninfected. Newly developed assays10 are more sensitive for detecting early infection; therefore the number of specimens collected in settings with high HIV incidence that will be negative by WB or IFA and require NAAT testing is expected to increase. The proposed algorithm, which utilizes a rapid supplemental test which is less sensitive for early infection than the screening test,2 will identify more persons in the early stage of infection than would be possible if the screening IA and rapid test had similar seroconversion sensitivities.11 The algorithm would thereby allow for more accurate and rapid distinction of persons with early infection, who are likely to be at high risk for HIV transmission, from persons with undiagnosed infection with lower transmission risk.12,13 In this study, all the screening IAs had similar performance, and therefore other characteristics that we did not evaluate, such as convenience, time-to-results and cost may be important in determining which test methods will be used in different HIV testing settings. For example, both the VITROS and ADVIA assays are run on automated analyzers that allow for loading of individual specimens, are commonly used in healthcare settings to perform a variety of clinical chemistry and serology tests, and provide results in less than an hour. However, use of these analyzers may be cost-prohibitive in some settings due to the costs associated with running appropriate controls for small patient specimen batch sizes. Even after allowing for repeat testing of screening IAs as
K.P. Delaney et al. / Journal of Clinical Virology 52S (2011) S5–S10
specified by manufacturers, using rapid HIV tests after obtaining reactive IA results may expedite the algorithm used to conduct HIV testing and allow more timely confirmation of HIV infection in healthcare settings. Currently, many laboratories that perform HIV screening tests send repeatedly reactive specimens out to reference laboratories for WB testing, increasing the turnaround time for a definitive test result. The proposed algorithm would allow on-site supplemental testing and may thereby obviate some of the challenges to implementation of routine testing for HIV in healthcare setting as recommended by CDC.14 The algorithm proposed at the 2010 HIV Diagnostics Conference5 suggested using an HIV-1/HIV-2 differentiation assay after a reactive HIV-1/HIV-2 screening IA, and others have shown this to be an effective strategy.15–17 In this study we compared the performance of 6 FDA-approved rapid tests for this role. Our finding that all the rapid tests performed similarly as supplemental tests in the algorithm is not unexpected.7 Still, because of the clinical differences in the progression and treatment of HIV-2 disease18 and the lack of an FDA-approved HIV-2 supplemental assay,15 use of an HIV-1/HIV-2 differentiation assay is likely to be the preferred approach. This study has several limitations. Insufficient specimen volumes prevented us from testing all specimens with all test methods. Of particular importance was the inability to subject all specimens with initially reactive IA results to repeat testing with IAs. Repeat testing of the ABBOTT assay had little effect on the number of specimens requiring NAAT. However, repeat testing of the BIORAD assay dramatically reduced the number of specimens with initial falsepositive results that would require a rapid test and NAAT. This is an important finding because it suggests that repeating IAs according to package insert instructions may be more cost-effective than subjecting these additional specimens to rapid testing and NAAT. In this study, we were unable to repeat testing on specimens with initially reactive ADVIA results, in accordance with package insert instructions, and therefore may have underestimated the performance of this test in the algorithm. In addition, the differences in total numbers of specimens tested using different rapid tests interfered with the ability to directly compare the number of NAAT tests required. Another limitation of this study is that none of the specimens collected were from HIV-2 infected individuals and so we could not evaluate how well the proposed algorithm performed at identifying HIV-2 infection. In summary, we found that, using specimens collected from persons at high risk for HIV infection, the proposed laboratory algorithm was able to identify nearly all true infections, including some acute infections and, when tests were performed according to their package insert instructions, produced no false-positive results. This alternative algorithm has the potential to identify both prevalent and acute infection with improved efficiency and fewer false-negative results than the current HIV testing algorithm used in the United States. Funding This work was funded by the Centers for Disease Control and Prevention. Competing interests None declared. Ethical approval The original study under which specimens were obtained from clients seeking HIV testing was reviewed and approved by both CDC
S9
and local institutional review boards. Study participants consented to storage and future testing of stored deidentified specimens, and this testing was subsequently conducted on review by the CDC after being categorized as research not involving identifiable human subjects. Disclaimer The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention or the U.S. Department of Health and Human Services or Quest Diagnostics, Inc. The use of trade names and commercial sources is for identification only and does not imply endorsement by the U.S. Department of Health and Human Services. Acknowledgements The authors would like to acknowledge study staff, especially Lashawnda Royal and Staeci Morita, as well as all the study participants who provided specimens for this study. We would like to acknowledge IA testing by Abbott Diagnostics, as well as the provision of APTIMA testing supplies by GenProbe. We would also like to thank Dollene Hemmerlein, Suzette Bartley, and the staff from the CDC serum bank for receiving, aliquoting and managing storage and shipment of study specimens. References 1. Centers for Disease Control and Prevention. Interpretation and use of the Western blot assay for serodiagnosis of human immunodeficiency virus type 1 infections. MMWR Recomm Rep 1989;38:S-7. 2. Owen SM, Yang C, Spira T, Ou Y, Pau CP, Parekh BS, et al. Alternative algorithms for human immunodeficiency virus infection diagnosis using tests that are licensed in the United States. J Clin Microbiol 2008;46(5): 1588–95. 3. Patel P, MacKellar D, Simmons P, Uniyal A, Gallager K, Bennett B, et al. Detecting acute human immunodeficiency virus infection using three different screening immunoassays and nucleic acid amplification testing for human immunodeficiency virus RNA 2006–2008. Arch Intern Med 2010;170(1): 66–74. 4. Stekler JD, Swenson PD, Coombs RW, Dragavon J, Thomas KK, Brennan CA, et al. HIV testing in a high-incidence population: is antibody testing alone good enough? Clin Infect Dis 2009;49:444–53. 5. Pandori MW, Branson BM. 2010 HIV diagnostics conference. Expert Rev Anti Infect Ther 2010;8(6):631–3. 6. CDC. FDA-approved rapid HIV antibody screening tests. Available from: [accessed http://www.cdc.gov/hiv/topics/testing/rapid/rt-comparison.htm 31.03.11]. 7. Delaney KP, Branson BM, Uniyal A, Phillips S, Candal D, Owen SM, et al. Evaluation of the performance characteristics of 6 rapid HIV antibody tests. Clin Infect Dis 2011;52(2):257–63. 8. Ethridge SF, Hart C, Hanson DL, Parker MM, Bennett B, et al. Performance of the APTIMA HIV-1 RNA qualitative assay with 16- and 32-member specimen pools. J Clin Microbiol 2010;48(9):3343–5. 9. Hall HI, Song R, Rhodes P, Prejean J, An Q, Lee LM, et al. Estimation of HIV incidence in the United States. JAMA 2008;300(5):520–9. 10. Chavez P, Wesolowski L, Patel P, Delaney K, Owen SM. Evaluation of the performance of the Abbott ARCHITECT HIV Ag/Ab Combo Assay. J Clinical Virol 2011;52(Suppl 1):S51–5. 11. Fiscus SA, Pilcher CD, Miller WC, Powers KA, Hoffman IF, Price M, et al. Rapid, real-time detection of acute HIV infection in patients in Africa. J Infect Dis 2007;195(3):416–24. 12. Wawer MJ, Gray RH, Sewankambo NK, Serwadda D, Li X, Laeyendecker O, et al. Rates of HIV-1 transmission per coital act, by stage of HIV-1 infection, in Rakai, Uganda. J Infect Dis 2005;191(May (9)):1403–9. 13. Hollingsworth TD, Anderson RM, Fraser C. HIV-1 transmission, by stage of infection. J Infect Dis 2008;198(September (5)):687–93. 14. CDC. Revised recommendations for HIV testing of adults, adolescents, and pregnant women in health-care settings. MMWR Recomm Rep 2006;55(RR14):1–17. 15. Nasrullah M, Ethridge SF, Delaney KP, Wesolowski LG, Granade TC, Schwendemann J, et al. Comparison of alternative interpretive criteria for the HIV-1
S10
K.P. Delaney et al. / Journal of Clinical Virology 52S (2011) S5–S10
Western blot and results of the Multispot HIV-1/HIV-2 Rapid Test for classifying HIV-1 and HIV-2 infections. J Clin Virol 2011;52(Suppl 1):S23–7. 16. Steyer LM, Sullivan TJ, Parker MM. Evaluation of an alternative supplemental testing strategy for HIV diagnosis by retrospective analysis of clinical HIV testing data. J Clin Virol 2011;52(Suppl 1):S35– 40.
17. Torian LV, Forgione LA, Punsalang AE, Pirillo RE, Oleszko WR. Comparison of Multispot EIA with Western blot for confirmatory serodiagnosis of HIV. J Clin Virol 2011;52(Suppl 1):S41–4. 18. Ntemgwa ML, d’Aquin Toni T, Brenner BG, Camacho RJ, Wainberg MA. Antiretroviral drug resistance in human immunodeficiency virus type 2. Antimicrob Agents Chemother 2009;53(9):3611–9.