Internet-recruited gay, bisexual, and other men who have sex with men (MSM) were offered HIV self-tests (HIVSTs) after completing baseline, 3-, 6-, and 9-month follow-up surveys. The surveys asked about the use and distribution of these HIVSTs. Among 995 who reported on their distribution of HIVSTs, 667 (67.0%) distributed HIVSTs to their social network associates (SNAs), which resulted in 34 newly identified HIV infections among 2301 SNAs (1.5%). The main reasons participants reported not distributing HIVSTs included: wanting to use the HIVSTs themselves (74.9%); thinking that their SNAs would get angry or upset if offered HIVSTs (12.5%); or not knowing that they could give the HIVSTs away (11.3%). Self-testing programs can provide multiple HIVSTs and encourage the distribution of HIVST by MSM to their SNAs to increase awareness of HIV status among persons disproportionately affected by HIV.

BACKGROUND: Geenius HIV 1/2 Supplemental Assay (Geenius; Bio-Rad Laboratories) is the only Food and Drug Administration-approved HIV-1/HIV-2 antibody differentiation test for the second step in the HIV laboratory testing algorithm. We characterized the occurrence of true HIV-1 and HIV-2 infections as well as false results in 6 US clinical laboratories using Geenius. METHODS: We examined routine HIV testing outcome data from the time the laboratories began using the algorithm with Geenius until September 30, 2017. We calculated the positive predictive value for Geenius HIV-1 and HIV-2 reactivity separately. RESULTS: Of 5,046,684 specimens tested, 41,791 had reactive antigen/antibody test results. Most specimens with reactive antigen/antibody results were HIV-1 antibody-positive established infections (n = 32,421), 1,865 of which also had indeterminate HIV-2 bands present. Ninety-three specimens were HIV-2 antibody positive or untypable for HIV-1/HIV-2 antibody. Acute HIV-1 infections were found in 528 specimens; 881 specimens lacked the nucleic acid test to determine the possibility of acute HIV-1 infection. False-positive antigen/antibody test results were present in 7505 specimens. Few specimens (n = 363) had false-positive antigen/antibody results with indeterminate Geenius and negative HIV-1 nucleic acid test results. The positive predictive values of Geenius reactivity were 99.4% for HIV-1 and 4.3% for HIV-2. CONCLUSIONS: Routine testing using the laboratory testing algorithm with Geenius resulted in most specimens resolving as HIV negative or HIV-1 positive. The occurrence of indeterminate HIV-2 bands with a Geenius final assay interpretation of HIV-1 positive was more common than true HIV-2 infections. Reporting indeterminate HIV-2 results in this situation may cause confusion with interpreting HIV infection status.

BACKGROUND: HIV testing guidelines provided by the Centers for Disease Control and Prevention (CDC) are continually changing to reflect advancements in new testing technology. Evaluation of existing and new point-of-care (POC) HIV tests is crucial to inform testing guidelines and provide information to clinicians and other HIV test providers. Characterizing the performance of POC HIV tests using unprocessed specimens can provide estimates for the window period of detection, or the time from HIV acquisition to test positivity, which allows clinicians and other HIV providers to select the appropriate POC HIV tests for persons who may be recently infected with HIV. OBJECTIVE: This paper describes the protocols and procedures used to evaluate the performance of the newest POC tests and determine their sensitivity during early HIV infection. METHODS: Project DETECT is a CDC-funded study that is evaluating POC HIV test performance. Part 1 is a cross-sectional, retrospective study comparing behavioral characteristics and HIV prevalence of the overall population of the Public Health-Seattle & King County (PHSKC) Sexually Transmitted Disease (STD) Clinic to Project DETECT participants enrolled in part 2. Part 2 is a cross-sectional, prospective study evaluating POC HIV tests in real time using unprocessed whole blood and oral fluid specimens. A POC nucleic acid test (NAT) was added to the panel of HIV tests in June 2018. Part 3 is a longitudinal, prospective study evaluating seroconversion sensitivity of POC HIV tests through serial follow-up testing. For comparison, HIV-1 RNA and HIV-1/HIV-2 antigen/antibody tests are also performed for participants enrolled in part 2 or 3. A behavioral survey that collects information about demographics, history of HIV testing, STD history, symptoms of acute HIV infection, substance use, sexual behaviors in the aggregate and with recent partners, and use of pre-exposure prophylaxis and antiretroviral therapy is completed at each part 2 or 3 visit. RESULTS: Between September 2015 and March 2019, there were 14,990 Project DETECT-eligible visits (part 1) to the PHSKC STD Clinic resulting in 1819 part 2 Project DETECT study visits. The longitudinal study within Project DETECT (part 3) enrolled 27 participants with discordant POC test results from their part 2 visit, and 10 (37%) were followed until they had fully seroconverted with concordant positive POC test results. Behavioral survey data and HIV test results, sensitivity, and specificity will be presented elsewhere. CONCLUSIONS: Studies such as Project DETECT are critical for evaluating POC HIV test devices as well as describing characteristics of persons at risk for HIV acquisition in the United States. HIV tests in development, including POC NATs, will provide new opportunities for HIV testing programs. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): RR1-10.2196/16332.

BACKGROUND: The performance of recently approved point-of-care (POC) HIV tests should be assessed using unprocessed specimens. OBJECTIVE: To evaluate the sensitivity and specificity of four POC HIV tests using whole blood (WB) and two using oral fluid (OF) among persons recruited from health clinics in Seattle, Washington, during September 2015-September 2017. STUDY DESIGN: Participants were tested with the POC tests, additional plasma and serum were collected for laboratory testing, and participant- reported use of antiretroviral therapy (ART) or pre-exposure prophylaxis (PrEP) was recorded. Participants testing negative on all tests could reenroll every 90 days. Specimens from persons previously diagnosed with HIV infection as well as from those who were newly diagnosed during the study were included in the sensitivity estimate. Sensitivity and specificity were calculated based on HIV status determined by laboratory testing. RESULTS: Of 1,256 visits, 179 were from persons with HIV infection; 120 of these were taking ART. Among 1,077 visits from participants not diagnosed with HIV, PrEP use was reported at 155 (14.4%) visits. Sensitivity was similar among POC WB tests (95.53%-97.21%; p>0.05). Among participants on ART, sensitivity was lower for the same test performed on OF compared to WB (p<0.003). Specificity was high for all tests (99.44%- 100.00%); we did not detect specificity differences with PrEP use. CONCLUSIONS: These POC tests displayed relatively high sensitivity and specificity using unprocessed specimens, suggesting their effectiveness in identifying HIV infections whenever laboratory-based testing is not feasible. Nonetheless, clients with recent risk should retest to rule out the possibility of a false-negative result.

BACKGROUND: There is benefit to early HIV-1 diagnosis and treatment, but there is no FDA-approved quantitative assay with a diagnostic claim. We compared the performance of the Hologic Aptima HIV-1 Quant (APT-Quant) and Aptima HIV-1 Qual (APT-Qual) assays for diagnostic use and the performance of a diagnostic algorithm consisting of Bio-Rad BioPlex 2200 HIV Ag-Ab assay (BPC) followed by APT-Quant (two-test) compare to BPC followed by Geenius HIV-1/2 supplemental assay (Geenius) with reflex to APT-Qual (three-test). METHODS: 524 plasma, which included 419 longitudinal specimens from HIV-1 seroconverters (78 were after initiating antiretroviral therapy (ART)) and 105 from ART-naive persons with established HIV-1 infections, were used to evaluate APT-Quant performance for diagnostic use. Specimens from 200 HIV-negative persons were used to measure specificity. For the algorithm comparison, BPC-reactive specimens were evaluated with the two-test or three-test algorithm. McNemar's test was used to compare performance. RESULTS: APT-Quant detected more samples early in infection compared with APT-Qual. APT-Quant specificity was 99.8%. Before ART initiation, the algorithms performed similarly among samples from different stages of infection. After ART initiation, the three-test algorithm performed significantly better (p=0.0233). CONCLUSIONS: APT-Quant has excellent performance for diagnostic use. The two-test algorithm works well in ART-naive samples, but its performance decreases after the IgG response is elicited and with ART-induced suppressed viremia. Providing confirmation and VL with one test result could be advantageous for patient care. However, additional factors and challenges associated with the implementation of this two-test algorithm such as cost, specimen type and collection need further evaluation.

Since 2014, the recommended laboratory testing algorithm for diagnosing human immunodeficiency virus (HIV) infection has included a supplemental HIV-1/HIV-2 differentiation test to confirm infection type on the basis of the presence of type-specific antibodies (1). Correctly identifying HIV-1 and HIV-2 infections is vital because their epidemiology and clinical management differ. To describe the percentage of diagnoses for which an HIV-1/HIV-2 differentiation test result was reported and to categorize HIV type based on laboratory test results, 2010-2017 data from CDC's National HIV Surveillance System (NHSS) were analyzed. During 2010-2017, a substantial increase in the number of HIV-1/HIV-2 differentiation test results were reported to NHSS, consistent with implementation of the HIV laboratory-based testing algorithm recommended in 2014. However, >99.9% of all HIV infections identified in the United States were categorized as HIV-1, and the number of HIV-2 diagnoses (mono-infection or dual-infection) remained extremely low (<0.03% of all HIV infections). In addition, the overall number of false positive HIV-2 test results produced by the HIV-1/HIV-2 differentiation increased. The diagnostic value of a confirmatory antibody differentiation test in a setting with sensitive and specific screening tests and few HIV-2 infections might be limited. Evaluation and consideration of other HIV tests approved by the Food and Drug Administration (FDA) that might increase efficiencies in the CDC and Association of Public Health Laboratories-recommended HIV testing algorithm are warranted.

BACKGROUND: In 2016, HIV-2 nucleic acid testing (NAT) was added to a shared service program that conducts HIV-1 NAT for public health laboratories performing the recommended algorithm for diagnosing HIV. Here we evaluate the usefulness of HIV-2 NAT in this program as compared to HIV-1 NAT. METHODS: Specimens eligible for HIV-1 NAT were reactive on an HIV-1/2 antibody or antigen/antibody initial test and non-reactive or indeterminate on a supplemental antibody test or were reactive for HIV-1 antigen-only on an HIV-1/2 antigen/antibody initial test. Specimens eligible for HIV-2 NAT were reactive on an initial test, HIV-2 indeterminate or HIV indeterminate on a supplemental antibody test and had no detectable HIV-1 RNA or were reactive for HIV-2 antibody on an HIV-1/2 antigen/antibody test and this reactivity was not confirmed with a supplemental antibody assay. All specimens were tested in a reference laboratory using APTIMA HIV-1 qualitative RNA and/or a validated qualitative HIV-2 RNA real-time PCR assay. RESULTS: During 2016-2019, HIV-1 RNA was detected in 234/1731 (14%) specimens tested. HIV-2 RNA was not detected in 52 specimens tested. Median time from specimen collection to reporting of HIV-1 and HIV-2 NAT results by year ranged from 9-10 days and 22-27 days, respectively. Two specimens with HIV-2 indeterminate results on a supplemental antibody test had detectable HIV-1 RNA. CONCLUSIONS: A shared service model for HIV-1 NAT is both feasible and beneficial for public health laboratories. However, because no HIV-2 infections were detected, our data suggest that this program should reconsider the usefulness of HIV-2 NAT testing.

BACKGROUND: Since 2014, the recommended algorithm for laboratory diagnosis of HIV infection in the United States has consisted of an HIV-1/2 antigen/antibody (Ag/Ab) test followed by an HIV-1/2 antibody (Ab) differentiation test and, if necessary, a diagnostic HIV-1 nucleic acid test (NAT) to resolve discordant or indeterminate results. METHODS: Using stored specimens from persons seeking HIV testing who had not received a previous diagnosis or treatment, we compared the performance of a three-step alternative algorithm consisting of an Ag/Ab test followed by a quantitative HIV-1 RNA viral load assay and, if viral load is not detected, an Ab differentiation test, to that of the recommended algorithm. We calculated the sensitivity and specificity of five Ag/Ab tests and the proportion of specimens correctly classified by the alternative algorithm compared to the recommended algorithm. Results were examined separately for specimens classified as early infection, established infection, and false-reactive screening RESULTS: Sensitivity and specificity were similar among all Ag/Ab tests. Viral load quantification correctly classified all specimens from early infection, all false-reactive screening specimens, and the majority of specimens from established infection. CONCLUSIONS: Although cost, regulatory barriers, test availability, and the ability to differentiate early from established infection must be considered, this alternative algorithm can potentially decrease the total number of tests performed and reduce turnaround time, thereby streamlining HIV diagnosis and initiation of treatment.

Since 2005, the HIV Diagnostics Conference has served as a central platform for fostering collaborations and partnerships among attendees who are involved in all aspects of HIV testing. The conference provides an open forum where attendees present and exchange ideas, review data on newer test technologies and algorithms, preview innovative testing methodologies and technologies, and present best practices related to testing, including how it relates to linkage to care, treatment, and prevention services. This approach has proven to be effective to encourage the advancement of HIV testing technology and strategies used in the US.

The Centers for Disease Control and Prevention recommends annual HIV tests for men who have sex with men (MSM), yet some have never tested. We analyzed data from the MSM Testing Initiative. Of 68,185 HIV tests, 8% were with MSM who never previously tested ("first-time testers"). Among tests with first-time testers, 70.7% were with MSM from racial or ethnic minorities; 66.5% were with MSM younger than 30 years. Tests with MSM who reported female partners only during the past year (compared to male partners only) or were recruited for at-home testing (compared to venue-based recruitment) were 4 times (prevalence ratio [PR] 3.62, 95% CI 3.15-4.15) and 5 times as likely (PR 4.69, 95% CI 4.22-5.21) to be associated with first-time testing. At-home testing and focusing on MSM who have sex with women may be effective methods for reaching MSM who are first-time testers.

BACKGROUND: The capacity of HIV Antigen/Antibody (Ag/Ab) immunoassays (IA) to detect HIV-1 p24 antigen has resulted in improved detection of HIV-1 infections in comparison to Ab-only screening assays. Since its introduction in the US, studies have shown that the Determine HIV-1/2 Ag/Ab Combo assay (Determine Ag/Ab) detects HIV infection earlier than laboratory-based IgM/IgG-sensitive IAs, but its sensitivity for HIV-1 p24 Ag detection is reduced compared to laboratory-based Ag/Ab assays. However, further evaluation is needed to assess its capacity to detect acute HIV-1 infection. OBJECTIVE: To assess the performance of Determine Ag/Ab in serum from acute HIV-1 infections. STUDY DESIGN: Select serum specimens that screened reactive on a laboratory-based Ag/Ab IA or IgM/IgG Ab-only IA, with a negative or indeterminate supplemental antibody test and detectable HIV-1 RNA were retrospectively tested with Determine Ag/Ab. Results were compared with those of the primary screening immunoassay to evaluate concordance within this set of algorithm-defined acute infections. RESULTS: Of 159 algorithm-defined acute HIV-1 specimens, Determine Ag/Ab was reactive for 105 resulting in 66.0% concordance. Of 125 that were initially detected by a laboratory-based Ag/Ab IA, 81 (64.8%) were reactive by Determine Ag/Ab. A total of 34 acute specimens were initially detected by a laboratory-based IgM/IgG Ab-only IA and 24 (70.6%) of those were reactive by Determine Ag/Ab. CONCLUSIONS: Due to their enhanced sensitivity, laboratory-based Ag/Ab IAs continue to be preferred over the Determine Ag/Ab as the screening method used by laboratories conducting HIV diagnostic testing on serum and plasma specimens.

HIV testing is a key component of HIV prevention. It is this critical clinical encounter that serves as the starting point for diagnosing and treating persons who are infected and delivering preventive services to those who are uninfected. Because HIV testing is so important to prevention strategies for controlling the HIV epidemic in the United States, we read with great interest the article by Hurt and colleagues1 in this issue, which provides an excellent overview of the current options available for HIV testing in clinical, nonclinical, and research settings. Their update highlights recent changes to nomenclature, updated data—particularly on the window period of HIV tests—and updates to the laboratory algorithm for diagnosis of HIV infection, at a time when this information is changing rapidly. | Hurt et al. refer to changes in the “official nomenclature” of HIV tests. Although the Centers for Disease Control and Prevention (CDC) does not determine official nomenclature for HIV test types, the CDC Division of HIV/AIDS Prevention has recently made changes to Web sites and other documents that refer to the different types of HIV tests. As discussed at the 2016 HIV Diagnostics Conference,2,3 the term “generations” began to appear in the literature shortly after HIV tests that used recombinant peptides instead of viral lysate antigens (the “2nd generation”) were developed.4–6 However, the “official” nomenclature likely gained traction when Owen et al.7 published an article including a discussion of generations, and CDC and others largely adopted the term for use in presentations, Web pages, and other documents. Indeed, a complete description of test generations appears in both the updated Clinical & Laboratory Standards Institute standards8 and the CDC/Association of Public Health Laboratories (APHL) guidelines for the laboratory diagnosis of HIV infection.9 However, as new HIV tests continued to become available, the lines between generations began to blur. In the 2008 article,7 the term generation was reserved for laboratory-based, instrumented immunoassays. As Hurt et al. reviewed, single-use, point-of-care rapid tests use different technology and probably should be considered separately. Nevertheless, both test manufacturers and authors evaluating these tests began to use the term generations to describe rapid tests. Originally, the generations described incremental improvements in test sensitivity and specificity. However, some of the newer tests within the same generation have different sensitivity for early infection.10 These differences can largely be explained by other aspects of test design, for example, whether they are lateral flow or immunconcentrating rapid tests, reagents used for detection of analytes, or the volume of sample required to perform the test.1 In addition, there are also IgG-sensitive rapid tests that differentiate HIV-1 from HIV-2, and new tests that differentiate p24-antigen detection from antibody detection, but have the same sensitivity during early infection as tests that report only one signal as “reactive for p24-antigen and/or HIV antibody.”1,3,10 As a result, in the article documenting seroconversion sensitivity on plasma specimens that Hurt et al. referenced,10 tests were described in terms of the analytes they can detect and the types of technology (instrumented, laboratory-based, vs. single-use, rapid) that they use to do so. These changes have been implemented in CDC Web pages and documents contained therein.11 In particular, the advantages/disadvantages of Food and Drug Administration–approved HIV tests guide12 may be particularly useful for clinicians and others who need to understand differences in characteristics of the tests available in the United States.

Identifying patients at-risk for HIV infection, such as men who have sex with men (MSM), is an important step in providing HIV testing and prevention interventions. It is unknown how primary care providers (PCPs) assess MSM status and related HIV-risk factors. We analyzed data from a panel-derived web-based survey for healthcare providers conducted in 2014 to describe how PCPs in the U.S. determined their patients' MSM status. We calculated adjusted prevalence ratios (aPR) and 95% confidence intervals (CI) to describe PCP characteristics associated with systematically determining MSM status (i.e., PCP used "a patient-completed questionnaire" or "routine verbal review of sex history"). Among the 1008 PCPs, 56% determined MSM status by routine verbal review of sexual history; 41% by patient disclosure; 39% by questions driven by symptoms/history; 23% by using a patient-completed questionnaire, and 9% didn't determine MSM status. PCPs who systematically determined MSM status (n=665; 66%) were more likely to be female (aPR=1.16, CI=1.06-1.26), to be affiliated with a teaching hospital (aPR=1.15, CI=1.06-1.25), to routinely screen all patients aged 13-64 for HIV (aPR=1.29, CI=1.18-1.41), and to estimate that 6% or more of their male patients are MSM (aPR=1.14, CI=1.01-1.30). The majority of PCPs assessed MSM status and HIV risk factors through routine verbal reviews of sexual history. Implementing a systematic approach to identify MSM status and assess risk may allow PCPs to identify more patients needing frequent HIV testing and other preventive services, while mitigating socio-cultural barriers to obtaining such information.

In the United States, an estimated 67% of new HIV diagnoses are among men who have sex with men (MSM), however 25% of HIV-positive MSM in the 2014 National HIV Behavioral Surveillance Survey were unaware of their infection. HIV self-testing (HIVST) with rapid diagnostic tests (RDTs) may facilitate access to HIV testing. We evaluated the ability of 22 MSM to conduct two HIV RDTs (OraQuick (R) In-Home HIV Test and a home-use prototype of Sure Check (R) HIV 1/2 Assay), interpret sample images of test results, and collect a dried blood spot (DBS) specimen. While some participants did not follow every direction, most participants were able to conduct HIVST and correctly interpret their results. Interpretation of panels of RDT images was especially difficult when the "control" line was missing, and 27% of DBS cards produced were rated as of bad quality. Modifications to the DBS instructions were necessary prior to evaluating the performance of these tests in real-world settings.

The 2016 HIV Diagnostics Conference, held in Atlanta, Georgia, was attended by public health officials, laboratorians, HIV testing program managers, surveillance coordinators and industry representatives. The conference addressed test performance data, the implementation of new testing algorithms, quality assurance, and the application of new tests in a variety of settings. With regard to the recommended Centers for Disease Control and Prevention/Association of Public Health Laboratories HIV laboratory testing algorithm, the conference featured performance data, implementation challenges such as a lack of test options for the second and third steps, as well as data needs for new tests that may be used as part of the algorithm. There are delays when nucleic acid testing is needed with the algorithm. Novel tests such as point of care nucleic acid tests are needed on the U.S. market to readily identify acute infection. Multiplex tests are being developed which allow for the simultaneous detection of multiple pathogens. CDC staff highlighted new guidance for testing in non-clinical settings. Innovative approaches to linking testing and care in some settings have led to identification of early infections, improved receipt of test results and expedited initiation of therapy. Work continues to optimize testing so that infections are accurately identified as early as possible and time to treatment is minimized to improve health outcomes and prevent transmission.

OBJECTIVE: In three U.S. State Public Health Laboratories (PHLs) using a fourth-generation immunoassay (IA), an HIV-1/HIV-2 differentiation antibody IA and a nucleic acid test (NAT), we characterized the yield and time to reporting of acute infections, and cost per positive specimen. METHODS: Routine HIV testing data were collected from July 1, 2012-June 30, 2013 for Massachusetts and Maryland PHLs, and from November 27, 2012-June 30, 2013 for Michigan PHL. Massachusetts and Michigan used fourth-generation and differentiation IAs with NAT conducted by a referral laboratory. In Maryland, fourth-generation IA repeatedly reactive specimens were followed by a Western blot (WB), and those with negative or indeterminate results were tested with a differentiation IA and HIV-1 NAT, and if positive by NAT, confirmed by a different HIV-1 NAT. Specimens from WB-positive persons at risk for HIV-2 were tested with a differentiation IA and, if positive, with an HIV-2 WB and/or differential HIV-1/HIV-2 proviral DNA polymerase chain reaction. RESULTS: Among 7914 specimens from Massachusetts PHL, 6069 from Michigan PHL, and 36,266 from Maryland PHL, 0.10%, 0.02% and 0.05% acute infections were identified, respectively. Massachusetts and Maryland PHLs each had 1 HIV-2 positive specimen. The median time from specimen receipt to laboratory reporting of results for acute infections at Massachusetts, Michigan and Maryland PHLs was 8, 11, and 7 days respectively. The laboratory cost per HIV positive specimen was $336 (Massachusetts), $263 (Michigan) and $210 (Maryland). CONCLUSIONS: Acute and established infections were found by PHLs using fourth-generation IA in conjunction with antibody tests and NAT. Time to reporting of acute HIV test results to clients was suboptimal, and needs to be streamlined to expedite treatment and interrupt transmission.

BACKGROUND: To improve clinical and public health outcomes through early human immunodeficiency virus (HIV) detection, fourth-generation antigen/antibody immunoassay (4IA) and supplemental testing results must be returned rapidly. METHODS: We examined HIV testing data at Harborview Medical Center (HMC), Massachusetts General Hospital (MGH), and the Medical University of South Carolina (MUSC), which used 4IA and supplemental antibody and nucleic acid tests (NATs). At MGH and MUSC, HIV-1 Western blot (WB) and HIV-2 testing were conducted at a reference laboratory. We compared time from specimen collection to laboratory result for established (positive WB) and acute infections (reactive 4IA, negative/indeterminate WB, detectable NAT), and we calculated testing cost per positive-test result. RESULTS: From 3731 (MUSC) to 19 774 (MGH) tests were conducted; 0.01% (MGH) to 0.05% (HMC) were acute infections. Each laboratory had reactive 4IA, WB-negative, or indeterminate specimens without NAT (ie, potential acute infections). Time to result was 1.5 (HMC) to 5.2 days (MGH) for acute and 1.0 (HMC) to 5.2 days (MGH) for established infections. Costs were $1054 (MGH) to $1521 (MUSC). CONCLUSIONS: Conducting supplemental testing in-house lowered turnaround times, which may be further reduced with rapid HIV-1/HIV-2 differentiation tests. Hospitals may benefit from quantitative NATs not requiring physician orders, so all potential acute infections receive NAT.

BACKGROUND: Many public health laboratories adopting the U.S. HIV laboratory testing algorithm do not have a nucleic acid test (NAT), which is needed when the third- or fourth-generation HIV screening immunoassay is reactive and the antibody-based supplemental test is non-reactive or indeterminate. OBJECTIVES: Among public health laboratories utilizing public health referral laboratories for NAT conducted as part of the algorithm, we evaluated the percentage of screening immunoassays needing NAT, the number of specimens not meeting APTIMA (NAT) specifications, time to APTIMA result, the proportion of acute infections (i.e., reactive APTIMA) among total infections, and screening immunoassay specificity. STUDY DESIGN: From August 2012 to April 2013, 22 laboratories enrolled to receive free APTIMA (NAT) at New York or Florida public health referral laboratories. Data were analyzed for testing conducted until June 2013. RESULTS: Submitting laboratories conducted a median of 4778 screening immunoassays; 0-1.3% (median 0.2%) needed NAT. Of 140 specimens received, 9 (6.4%) did not meet NAT specifications. The median time from specimen collection to reporting the 11 reactive NAT results was ten days, including six days from receipt in the submitting laboratory to shipment to the referral laboratory. Acute infections ranged from 0 to 12.5% (median 0%) of total infections. Third- and fourth-generation immunoassays met package insert specificity values. CONCLUSIONS: Public health referral laboratories provide a feasible option for conducting NAT. Reducing the time from specimen collection to submission of specimens for NAT is an important step toward maximizing the public health impact of identifying acute infections.

BACKGROUND: The use of Western blot (WB) as a supplemental test after reactive sensitive initial assays can lead to inconclusive or misclassified HIV test results, delaying diagnosis. OBJECTIVE: To determine the proportion of specimens reactive by immunoassay (IA) but indeterminate or negative by WB that could be resolved by alternative supplemental tests recommended under a new HIV diagnostic testing algorithm. STUDY DESIGN: Remnant HIV diagnostic specimens that were reactive on 3rd generation HIV-1/2 IA and either negative or indeterminate by HIV-1 WB from 11 health departments were tested with the Bio-Rad Multispot HIV-1/HIV-2 Rapid Test (Multispot) and the Gen-Probe APTIMA HIV-1 RNA Qualitative Assay (APTIMA). RESULTS: According to the new testing algorithm, 512 (89.8%) specimens were HIV-negative, 55 (9.6%) were HIV-1 positive (including 19 [3.3%] that were acute HIV-1 and 9 [1.6%] that were positive for HIV-1 by Multispot but APTIMA-negative), 2 (0.4%) were HIV-2 positive, and 1 (0.2%) was HIV-positive, type undifferentiated. 47 (21.4%) of the 220 WB-indeterminate and 8 (2.3%) of the 350 WB-negative specimens were HIV-1 positive. CONCLUSION: Applying the new HIV diagnostic algorithm retrospectively to WB-negative and indeterminate specimens, the HIV infection status could be established for nearly all of the specimens. IA-reactive HIV-infected persons with WB-negative results had been previously misclassified as uninfected, and HIV diagnosis was delayed for those with WB-indeterminate specimens. These findings underscore the limitations of the WB to confirm HIV infection after reactive results from contemporary 3rd or 4th generation IAs that can detect HIV antibodies several weeks sooner than the WB.

BACKGROUND: An alternative HIV testing algorithm, designed to improve the detection of acute and early infections and differentiate between HIV-1 and HIV-2 antibodies, has been developed by the Centers for Disease Control and Prevention and the Association of Public Health Laboratories. While it promises greater sensitivity, it also raises concerns about costs. OBJECTIVE: We sought to compare the most commonly used algorithm which was developed in 1989, a third-generation (3G) immunoassay (IA) and Western blot confirmatory test, to a newer algorithm. The new algorithm includes either a 3G or a fourth-generation (4G) initial IA, followed by confirmatory testing with a HIV-1/HIV-2 differentiation IA and, if needed, a nucleic acid amplification test (NAT). STUDY DESIGN: We conducted an analysis of HIV testing costs from the perspective of the laboratory, and classified costs according to IA testing volume. We developed a decision analytic model, populated with cost data from 17 laboratories and published assay performance data, to compare the cost-effectiveness of the testing algorithms for a cohort of 30,000 specimens with a 1% HIV prevalence and 0.1% acute HIV infection prevalence. RESULTS: Costs were lower in high-volume laboratories regardless of testing algorithm. For specimens confirmed positive for HIV antibody, the alternative algorithm (IA, Multispot) was less costly than the current algorithm (IA, WB); however, there was wide variation in reported testing costs. For our cohort, the alternative algorithm initiated with a 3G IA and 4G IA identified 15 and 25 more HIV infections, respectively, than the 1989 algorithm. In medium-volume laboratories, the 1989 algorithm was more costly and less effective than the alternative algorithm with a 3G IA; in high-volume laboratories, the alternative algorithm with 3G IA costs $162 more per infection detected. The alternative algorithm with 4G instead of 3G incurred an additional cost of $14,400 and $4865 in medium- and high-volume labs, respectively. DISCUSSION: HIV testing costs varied with IA testing volumes. The additional cost of 4G over 3G IA might be justified by the additional cases of HIV detected and transmissions averted due to earlier detection. CONCLUSION: The alternative HIV testing algorithm compares favorably to the 1989 algorithm in terms of cost and effectiveness.

BACKGROUND: An alternate HIV testing algorithm has been proposed which includes a fourth-generation immunoassay followed by an HIV-1/HIV-2 antibody differentiation supplemental test for reactive specimens and a nucleic acid test (NAT) for specimens with discordant results. OBJECTIVE: To evaluate the performance of five rapid tests (Alere Clearview, Bio-Rad Multispot, OraSure OraQuick, MedMira Reveal, and Trinity Biotech Unigold) as the supplemental antibody assay in the algorithm. STUDY DESIGN: A total of 3273 serum and plasma specimens that were third-generation immunoassay repeatedly reactive and Western blot (WB) negative or indeterminate were tested with rapid tests and NAT. Specimens were classified by NAT: (1) HIV-1 infected (NAT-reactive; n=184, 5.6%), (2) HIV-status unknown (NAT nonreactive; n=3078, 94.2%) or by Multispot, (3) HIV-2 positive (n=5), and (4) HIV-1 and HIV-2 positive (n=6). Excluding HIV-2 positive specimens, we calculated the proportion of reactive rapid tests among specimens with reactive and nonreactive NAT. RESULTS: The proportion of infected specimens with reactive rapid test results and negative or indeterminate WB ranged from 30.4% (56) to 47.8% (88) depending on the rapid test. From 1% to 2% of NAT-negative specimens had reactive rapid test results. CONCLUSIONS: In these diagnostically challenging specimens, all rapid tests identified infections that were missed by the Western blot, but only Multispot could differentiate HIV-1 from HIV-2. Regardless of which rapid test is used as a supplemental test in the alternative algorithm, false-positive algorithm results (i.e., reactive screening and rapid test in uninfected person) may occur, which will need to be resolved during the baseline medical evaluation.

OBJECTIVE: We evaluated the performance of the GS fourth-generation antigen/antibody assay and compared CDC's proposed alternative algorithm (repeatedly reactive [RR] fourth-generation immunoassay [IA] followed by an HIV-1/HIV-2 differentiation IA and, if needed, nucleic acid testing [NAT]) with the current algorithm (RR third-generation IA followed by HIV-1 Western blot [WB]). DESIGN: A convenience sample of the following four specimen sets was acquired: 10 014 from insurance applicants, 493 known WB-positive, 20 known WB-indeterminate specimens, and 230 specimens from 26 HIV-1 seroconverters. METHODS: Specimens were tested with the GS third- and fourth-generation IAs, the Multispot HIV-1/HIV-2 differentiation IA, NAT, and WB. We applied the two algorithms using these results. RESULTS: Among insurance specimens, 13 (0.13%) specimens were IA RR: 2 were HIV-positive (RR by third- and fourth-generation IAs, and WB and Multispot positive); 2 third-generation RR and 9 fourth-generation RR specimens were false-positive. Third- and fourth-generation specificities were 99.98% (95%CI: 99.93%-100%) and 99.91% (95%CI: 99.84%-99.96%) respectively.All HIV-1 WB-positive specimens were RR by third- and fourth-generation IAs. By Multispot, 491 (99.6%) were HIV-1 positive and 2 (0.4%) were HIV-2 positive. Only eight (40%) WB-indeterminate specimens were fourth-generation RR: 6 were Multispot and NAT negative and 2 were Multispot HIV-1 positive but NAT negative.The alternative algorithm correctly classified as positive 102 seroconverter specimens with the third-generation IA and 130 with the fourth-generation IA compared with 56 using the WB with either IA. CONCLUSIONS: The alternative testing algorithm improved early infection sensitivity and identified HIV-2 infections. Two potential false-positive algorithm results occurred with WB-indeterminate specimens.

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((R))), 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.

BACKGROUND: Blood specimens for HIV testing are frequently collected using serum collection tubes. The Aptima HIV-1 RNA Qualitative test, originally approved for diagnostic use as a supplemental test for use with plasma, is now approved for use with serum, and may be used in new laboratory-based HIV testing algorithms which detect acute and established HIV infections. OBJECTIVES: To compare the sensitivity of Aptima using serum and plasma specimens from persons with established HIV infection. STUDY DESIGN: Parallel serum and plasma specimens were collected from 325 persons with established HIV-1 infection who had positive immunoassay (IA) and Western blot (WB) results. Samples with negative Aptima results were considered false-negative and were subjected to repeat testing. Aptima sensitivity for serum and plasma was calculated relative to IA and WB, and compared using the McNemar test. RESULTS: The sensitivity of Aptima using serum (97.23%, 95% confidence interval [CI] 94.81-98.73) was similar to that using plasma (97.54%, 95% [CI] 95.21-98.93) p=1.00. Five of ten specimens initially false-negative on either serum or plasma were reactive on repeat testing. No specimens initially classified as false-negative on both matrices were reactive on both matrices on repeat testing. CONCLUSIONS: In specimens from persons with established infections, Aptima performed with similar sensitivity when used with serum or plasma. Using serum for immunoassay screening and supplemental testing may provide added convenience for laboratories.

BACKGROUND: The HIV-1 Western blot (WB) and immunofluorescence assay used to confirm HIV infections are less sensitive during seroconversion than immunoassays (IAs) used for screening. An alternative diagnostic algorithm has been proposed to detect early HIV-1 infection and differentiate HIV-1 from HIV-2. OBJECTIVES: We evaluated the performance of an algorithm with a third generation IA that when reactive was followed by a rapid test (Multispot) that differentiates HIV-1 from HIV-2. Multispot-reactive specimens were considered HIV-infected. Multispot-negative specimens were tested with a nucleic acid amplification test (NAAT) for resolution. STUDY DESIGN: WB-positive specimens [serum (n=2202), plasma (n=1109) and peripheral blood mononuclear cells (PBMCs) (n=1065)] were obtained from HIV-infected persons not taking antiretrovirals. HIV-uninfected specimens [plasma (n=1517) and PBMCs (n=1508)] with negative IA and NAAT results were obtained from blood donors. Specimens were tested with third generation IAs (Abbott rDNA, ADVIA Centaur, GS HIV1-2 Plus O, Ortho VITROS) in singlet, Multispot, and NAAT (APTIMA (RNA) and AMPLICOR (DNA)). We calculated algorithm sensitivity and specificity and the proportion of IA-reactive specimens requiring NAAT. RESULTS: Algorithm sensitivity was 99.95% with APTIMA and 100% with AMPLICOR. One WB-positive specimen reactive by all IAs and AMPLICOR was negative by Multispot and APTIMA. Algorithm specificity was 100% using APTIMA or AMPLICOR as NAAT. From 0.10% (Abbott) to 2.43% (VITROS) of IA-reactive specimens required NAAT. CONCLUSIONS: The proposed algorithm performs with high sensitivity and specificity in specimens from persons with established HIV infection and uninfected blood donors and appears to be a good alternative to the current algorithm.

BACKGROUND: HIV-1 Western blot (WB) may be positive in specimens from persons with HIV-2 infection due to cross-reactive antibodies. HIV-1 and HIV-2 infections may be identified using assays designed to differentiate HIV-1 and HIV-2 antibody reactivity. OBJECTIVES: To evaluate the ability of the current CDC WB criteria, alternative more stringent HIV-1 WB criteria (2 env plus one gag or pol band) and the Multispot HIV-1/HIV-2 Rapid Test to accurately differentiate HIV-1 and HIV-2 infections. STUDY DESIGN: Two panels were used to determine the ability of each method to properly classify HIV-1 and HIV-2 infections: an HIV-2 panel (n=114) determined to be HIV-2 antibody-positive by both Multispot and by a validated HIV-2 WB, and 2135 HIV-1/HIV-2 immunoassay repeatedly reactive (IA-RR) specimens from the New York State Department of Health Laboratory (NYS). RESULTS: By CDC WB criteria, 53 (46.5%) HIV-2 panel specimens were HIV-1 WB positive, 60 (52.6%) were indeterminate, and 1 (0.9%) was negative; the alternative WB criteria re-classified 75.5% of the positives as indeterminate. Among 2135 NYS IA-RR specimens, the alternative WB criteria increased the proportion of indeterminates by 0.8%. Only 6 (0.3%) of the NYS specimens were determined to be HIV-2 infections; all 6 were classified either as HIV-1 positive or indeterminate by both WB criteria, but were classified as HIV-2 (n=4) or HIV-1/2 undifferentiated (n=2) by Multispot. CONCLUSIONS: The alternative WB criteria classified most of the HIV-2 specimens that were HIV-1 positive by CDC criteria as indeterminate, but also slightly increased the proportion of HIV-1 specimens classified as indeterminate. The WB indeterminate specimens would require further testing or follow-up to resolve the infection status, whereas Multispot directly distinguished HIV-1 from HIV-2.

OBJECTIVE: Examine whether false-positive HIV enzyme immunoassay (EIA) test results occur more frequently among pregnant women than among women who are not pregnant and men (others). DESIGN: To obtain a large number of pregnant women and others tested for HIV, we identified specimens tested at a national laboratory using Genetic Systems HIV-1/HIV-2 Plus O EIA from July 2007 to June 2008. METHODS: Specimens with EIA repeatedly reactive and Western blot-negative or indeterminate results were considered EIA false-positive. We compared the false-positive rate among uninfected pregnant women and others, adjusting for HIV prevalence. Among all reactive EIAs, we evaluated the proportion of false-positives, positive predictive value (PPV), and Western blot bands among indeterminates, by pregnancy status. RESULTS: HIV prevalence was 0.06% among 921,438 pregnant women and 1.34% among 1,103,961 others. The false-positive rate was lower for pregnant women than others (0.14% vs. 0.21%, odds ratio 0.65 [95% confidence interval 0.61, 0.70]). Pregnant women with reactive EIAs were more likely than others (p<0.01) to have Western blot-negative (52.9% vs. 9.8%) and indeterminate results (17.0% vs. 3.7%) and lower PPV (30% vs. 87%). The p24 band was detected more often among pregnant women (p<0.01). CONCLUSIONS: False-positive HIV EIA results were rare and occurred less frequently among pregnant women than others. Pregnant women with reactive EIAs were more likely to have negative and indeterminate Western blot results due to lower HIV prevalence and higher p24 reactivity, respectively. Indeterminate results may complicate clinical management during pregnancy. Alternative methods are needed to rule out infection in persons with reactive EIAs from low prevalence populations.

The CDC recommends that a reactive rapid human immunodeficiency virus (HIV) test be confirmed with an approved supplemental test; the performance of an intermediate enzyme immunoassay (EIA) is optional. In support of this recommendation, it was found that of 1,431 reactive rapid HIV test results, 2 (0.1%) had false-negative oral fluid Western blot results and both had false-negative EIA results.

Rapid HIV testing is often conducted in non-clinical settings by staff with limited training, so quality assurance (QA) monitoring is critical to ensure accuracy of test results. Rapid tests (n=86,749) were generally conducted according to manufacturer instructions, but ongoing testing competency assessments and on-site QA monitoring were not uniformly conducted.