Multiassay Algorithm Research Articles

Development of a multiassay algorithm (MAA) to identify recent HIV infection in newly diagnosed individuals in Indonesia

Ongoing HIV transmission is a public health priority in Indonesia. We developed a new multiassay algorithm (MAA) to identify recent HIV infection. The MAA is a sequential decision tree based on multiple biomarkers, starting with CD4+ Tcells >200/μL, followed by plasma viral load (pVL)>1,000 copies/ml, avidity index (AI)<0 · 7, and pol ambiguity <0 · 47%. Plasma from 140 HIV-infected adults from 19 hospitals across Indonesia (January 2018- June 2020) was studied, consisting of a training set (N= 60) of longstanding infection (>12-month) and a test set (N= 80) of newly diagnosed (≤1-month) antiretroviral (ARV) drug naive individuals. Ten of eighty (12 · 5%) newly diagnosed individuals were classified as recent infections. Drug resistance mutations (DRMs) against reverse transcriptase inhibitors were identified in two individuals: one infected with HIV subtype C (K219Q, V179T) and the other with CRF01_AE (V179D). Ongoing HIV transmission, including infections with DRMs, is substantial in Indonesia.

Evaluation of multi-assay algorithms for cross-sectional HIV incidence estimation in settings with universal antiretroviral treatment

BackgroundMulti-assay algorithms (MAAs) are used to estimate population-level HIV incidence and identify individuals with recent infection. Many MAAs use low viral load (VL) as a biomarker for long-term infection. This could impact incidence estimates in settings with high rates of early HIV treatment initiation. We evaluated the performance of two MAAs that do not include VL.MethodsSamples were collected from 219 seroconverters (infected < 1 year) and 4376 non-seroconverters (infected > 1 year) in the HPTN 071 (PopART) trial; 28.8% of seroconverter samples and 73.2% of non-seroconverter samples had VLs ≤ 400 copies/mL. Samples were tested with the Limiting Antigen Avidity assay (LAg) and JHU BioRad-Avidity assays. Antibody reactivity to two HIV peptides was measured using the MSD U-PLEX assay. Two MAAs were evaluated that do not include VL: a MAA that includes the LAg-Avidity assay and BioRad-Avidity assay (LAg + BR) and a MAA that includes the LAg-Avidity assay and two peptide biomarkers (LAg + PepPair). Performance of these MAAs was compared to a widely used MAA that includes LAg and VL (LAg + VL).ResultsThe incidence estimate for LAg + VL (1.29%, 95% CI: 0.97–1.62) was close to the observed longitudinal incidence (1.34% 95% CI: 1.17–1.53). The incidence estimates for the other two MAAs were higher (LAg + BR: 2.56%, 95% CI 2.01–3.11; LAg + PepPair: 2.84%, 95% CI: 1.36–4.32). LAg + BR and LAg + PepPair also misclassified more individuals infected > 2 years as recently infected than LAg + VL (1.2% [42/3483 and 1.5% [51/3483], respectively, vs. 0.2% [6/3483]). LAg + BR classified more seroconverters as recently infected than LAg + VL or LAg + PepPair (80 vs. 58 and 50, respectively) and identified ~ 25% of virally suppressed seroconverters as recently infected.ConclusionsThe LAg + VL MAA produced a cross-sectional incidence estimate that was closer to the longitudinal estimate than two MAAs that did not include VL. The LAg + BR MAA classified the greatest number of individual seroconverters as recently infected but had a higher false recent rate.

Validation of population-level HIV-1 incidence estimation by cross-sectional incidence assays in the HPTN 071 (PopART) trial.

IntroductionCross‐sectional incidence testing is used to estimate population‐level HIV incidence and measure the impact of prevention interventions. There are limited data evaluating the accuracy of estimates in settings where antiretroviral therapy coverage and levels of viral suppression are high. Understanding cross‐sectional incidence estimates in these settings is important as viral suppression can lead to false recent test results. We compared the accuracy of multi‐assay algorithms (MAA) for incidence estimation to that observed in the community‐randomized HPTN 071 (PopART) trial, where the majority of participants with HIV infection were virally suppressed.MethodsHIV incidence was assessed during the second year of the study, and included only individuals who were tested for HIV at visits 1 and 2 years after the start of the study (2016–2017). Incidence estimates from three MAAs were compared to the observed incidence between years 1 and 2 (MAA‐C: LAg‐Avidity <2.8 ODn + BioRad Avidity Index <95% + VL >400 copies/ml; LAg+VL MAA: LAg‐Avidity <1.5 ODn + VL >1000 copies/ml; Rapid+VL MAA: Asanté recent rapid result + VL >1000 copies/ml). The mean duration of recent infection (MDRI) used for the three MAAs was 248, 130 and 180 days, respectively.Results and discussionThe study consisted of: 15,845 HIV‐negative individuals; 4406 HIV positive at both visits; and 221 who seroconverted between visits. Viral load (VL) data were available for all HIV‐positive participants at the 2‐year visit. Sixty four (29%) of the seroconverters and 3227 (72%) prevelant positive participants were virally supressed (<400 copies/ml). Observed HIV incidence was 1.34% (95% CI: 1.17–1.53). Estimates of incidence were similar to observed incidence for MAA‐C, 1.26% (95% CI: 1.02–1.51) and the LAg+VL MAA, 1.29 (95% CI: 0.97–1.62). Incidence estimated by the Rapid+VL MAA was significantly lower than observed incidence (0.92%, 95% CI: 0.69–1.15, p<0.01).ConclusionsMAA‐C and the LAg+VL MAA provided accurate point estimates of incidence in this cohort with high levels of viral suppression. The Rapid+VL significantly underestimated incidence, suggesting that the MDRI recommended by the manufacturer is too long or the assay is not accurately detecting enough recent infections.

HIV drug resistance in a cohort of HIV-infected MSM in the United States.

To analyze HIV drug resistance among MSM recruited for participation in the HPTN 078 study, which evaluated methods for achieving and maintaining viral suppression in HIV-infected MSM. Individuals were recruited at four study sites in the United States (Atlanta, Georgia; Baltimore, Maryland; Birmingham, Alabama; and Boston, Massachusetts; 2016-2017). HIV genotyping was performed using samples collected at study screening or enrollment. HIV drug resistance was evaluated using the Stanford v8.7 algorithm. A multiassay algorithm was used to identify individuals with recent HIV infection. Clustering of HIV sequences was evaluated using phylogenetic methods. High-level HIV drug resistance was detected in 44 (31%) of 142 individuals (Atlanta: 21%, Baltimore: 29%, Birmingham: 53%, Boston: 26%); 12% had multiclass resistance, 16% had resistance to tenofovir or emtricitabine, and 8% had resistance to integrase strand transfer inhibitors (INSTIs); 3% had intermediate-level resistance to second-generation INSTIs. In a multivariate model, self-report of ever having been on antiretroviral therapy (ART) was associated with resistance (P = 0.005). One of six recently infected individuals had drug resistance. Phylogenetic analysis identified five clusters of study sequences; two clusters had shared resistance mutations. High prevalence of drug resistance was observed among MSM. Some had multiclass resistance, resistance to drugs used for preexposure prophylaxis (PrEP), and INSTI resistance. These findings highlight the need for improved HIV care in this high-risk population, identification of alternative regimens for PrEP, and inclusion of integrase resistance testing when selecting ART regimens for MSM in the United States.

Targeting the HIV Epidemic in South Africa: The Need for Testing and Linkage to Care in Emergency Departments

BackgroundThe Eastern Cape province of South Africa has one of the highest burdens of HIV in the world. Emergency Departments (EDs) can serve as optimal clinical sites for the identification of new HIV infections and entry into care. We sought to determine the current burden of HIV disease among ED patients in the Eastern Cape. MethodsWe conducted a prospective cross-sectional observational study in the EDs of three Hospitals in the Eastern Cape province of South Africa from June 2017 to July 2018. All adult, non-critical patients presenting to the ED were systematically approached and offered a Point-Of-Care (POC) HIV test in accordance with South African guidelines. All HIV-positive individuals had their blood tested for the presence of antiretroviral therapy (ART) and the presence of viral suppression (≤1000 copies/ml). HIV incidence was estimated using a multi-assay algorithm, validated for a subtype C epidemic. FindingsOf the 2901 patients for whom HIV status was determined (either known HIV-positive or underwent POC HIV testing), 811 (28.0%) were HIV positive, of which 234 (28.9%) were newly diagnosed. HIV prevalence was higher in Mthatha [34% (388/1134) at Mthatha Regional Hospital and 28% (142/512) at Nelson Mandela Academic Hospital], compared to Port Elizabeth [22% (281/1255) at Livingstone Hospital]. HIV incidence was estimated at 4.5/100 person-years (95% CI: 2.4, 6.50) for women and 1.5 (CI 0.5, 2.5) for men. Of all HIV positive individuals tested for ART (585), 54% (316/585) tested positive for the presence of ARTs, and for all HIV positive participants with viral load data (609), 49% (299/609) were found to be virally suppressed. InterpretationOur study not only observed a high prevalence and incidence of HIV among ED patients but also highlights significant attrition along the HIV care cascade for HIV positive individuals. Furthermore, despite developing an optimal testing environment, we were only able to enrol a small sub-set of the ED population. Given the high HIV prevalence and high attrition in the ED population, HIV services in the ED should also develop strategies that can accommodate large testing volumes and ART initiation.

Cross-Sectional HIV Incidence Estimation with Missing Biomarkers.

Considerable progress has been made in the development of approaches for HIV incidence estimation based on a cross-sectional survey for biomarkers of recent infection. Multiple biomarkers when used in combination can increase the precision of cross-sectional HIV incidence estimates. Multi-assay algorithms (MAAs) for cross-sectional HIV incidence estimation are hierarchical stepwise algorithms for testing the biological samples with multiple biomarkers. The objective of this paper is to consider some of the statistical challenges for addressing the problem of missing biomarkers in such testing algorithms. We consider several methods for handling missing biomarkers for (1) estimating the mean window period, and (2) estimating HIV incidence from a cross sectional survey once the mean window period has been determined. We develop a conditional estimation approach for addressing the missing data challenges and compare that method with two naïve approaches. Using MAAs developed for HIV subtype B, we evaluate the methods by simulation. We show that the two naïve estimation methods lead to biased results in most of the missing data scenarios considered. The proposed conditional approach protects against bias in all of the scenarios.

Comparison of Self-report to Biomarkers of Recent HIV Infection: Findings from the START Trial.

Identifying individuals with recent HIV infection is critical to research related to viral reservoirs, outbreak investigations and intervention applications. A multi-assay algorithm (MAA) for recency of infection was used in conjunction with self-reported date of infection and documented date of diagnosis to estimate the number of participants recently infected in the Strategic Timing of AntiRetroviral Treatment (START) trial. We tested samples for three groups of participants from START using a MAA: (1) 167 individuals who reported being infected≤6months before randomization; (2) 771 individuals who did not know their date of infection but were diagnosed within 6months before randomization; and (3) as controls for the MAA, 199 individuals diagnosed with HIV≥2years before randomization. Participants with low titer and avidity and a baseline viral load>400 copies/mL were classified as recently infected. A significantly higher percentage of participants who self-reported being infected≤6months were classified as recently infected compared to participants diagnosed≥2years (65% [109/167] vs. 2.5% [5/199], p<0.001). Among the 771 individuals who did not know their duration of infection at randomization, 206 (26.7%) were classified as recently infected. Among those diagnosed with HIV in the 6months prior to enrollment, the 373 participants who reported recent infection (n=167) or who had confirmed recent infection by the MAA (n=206) differed significantly on a number of baseline characteristics from those who had an unknown date of infection and were not confirmed by the MAA (n=565). Participants recently infected by self-report and/or MAA were younger, more likely to be Asian, less likely to be black, less likely to be heterosexual, more likely to be enrolled at sites in the U.S., Europe or Australia, and have higher HIV RNA levels. There was good agreement between self-report of recency of infection and the MAA. We estimate that 373 participants enrolled in START were infected within 6months of randomization. Compared to those not recently infected, these participants were younger, had higher HIV RNA levels and were more likely to come from high income countries and from populations such as MSM with more regular HIV testing.

Identification and validation of a multi-assay algorithm for cross-sectional HIV incidence estimation in populations with subtype C infection.

IntroductionCross‐sectional methods can be used to estimate HIV incidence for surveillance and prevention studies. We evaluated assays and multi‐assay algorithms (MAAs) for incidence estimation in subtype C settings.MethodsWe analysed samples from individuals with subtype C infection with known duration of infection (2442 samples from 278 adults; 0.1 to 9.9 years after seroconversion). MAAs included 1‐4 of the following assays: Limiting Antigen Avidity assay (LAg‐Avidity), BioRad‐Avidity assay, CD4 cell count and viral load (VL). We evaluated 23,400 MAAs with different assays and assay cutoffs. We identified the MAA with the largest mean window period, where the upper 95% confidence interval (CI) of the shadow was <1 year. This MAA was compared to the LAg‐Avidity and BioRad‐Avidity assays alone, a widely used LAg algorithm (LAg‐Avidity <1.5 OD‐n + VL >1000 copies/mL), and two MAAs previously optimized for subtype B settings. We compared these cross‐sectional incidence estimates to observed incidence in an independent longitudinal cohort.ResultsThe optimal MAA was LAg‐Avidity <2.8 OD‐n + BioRad‐Avidity <95% + VL >400 copies/mL. This MAA had a mean window period of 248 days (95% CI: 218, 284), a shadow of 306 days (95% CI: 255, 359), and provided the most accurate and precise incidence estimate for the independent cohort. The widely used LAg algorithm had a shorter mean window period (142 days, 95% CI: 118, 167), a longer shadow (410 days, 95% CI; 318, 491), and a less accurate and precise incidence estimate for the independent cohort.ConclusionsAn optimal MAA was identified for cross‐sectional HIV incidence in subtype C settings. The performance of this MAA is superior to a testing algorithm currently used for global HIV surveillance.

Analysis of HIV Diversity in HIV-Infected Black Men Who Have Sex with Men (HPTN 061).

BackgroundHIV populations often diversify in response to selective pressures, such as the immune response and antiretroviral drug use. We analyzed HIV diversity in Black men who have sex with men who were enrolled in the HIV Prevention Trials Network 061 study.MethodsA high resolution melting (HRM) diversity assay was used to measure diversity in six regions of the HIV genome: two in gag, one in pol, and three in env. HIV diversity was analyzed for 146 men who were HIV infected at study enrollment, including three with acute infection and 13 with recent infection (identified using a multi-assay algorithm), and for 21 men who seroconverted during the study. HIV diversification was analyzed in a paired analysis for 62 HIV-infected men using plasma samples from the enrollment and 12-month (end of study) visits.ResultsMen with acute or recent infection at enrollment and seroconverters had lower median HRM scores (lower HIV diversity) than men with non-recent infection in all six regions analyzed. In univariate analyses, younger age, higher CD4 cell count, and HIV drug resistance were associated with lower median HRM scores in multiple regions; ARV drug detection was marginally associated with lower diversity in the pol region. In multivariate analysis, acute or recent infection (all six regions) and HIV drug resistance (both gag regions) were associated with lower median HRM scores. Diversification in the pol region over 12 months was greater for men with acute or recent infection, higher CD4 cell count, and lower HIV viral load at study enrollment.ConclusionsHIV diversity was significantly associated with duration of HIV infection, and lower gag diversity was observed in men who had HIV drug resistance. HIV pol diversification was more pronounced in men with acute or recent infection, higher CD4 cell count, and lower HIV viral load.

Viral and Host Characteristics of Recent and Established HIV-1 Infections in Kisumu based on a Multiassay Approach.

Integrated approaches provide better understanding of HIV/AIDS epidemics. We optimised a multiassay algorithm (MAA) and assessed HIV incidence, correlates of recent infections, viral diversity, plus transmission clusters among participants screened for Kisumu Incidence Cohort Study (KICoS1) (2007–2009). We performed BED-CEIA, Limiting antigen (LAg) avidity, Biorad avidity, and viral load (VL) tests on HIV-positive samples. Genotypic analyses focused on HIV-1 pol gene. Correlates of testing recent by MAA were assessed using logistic regression model. Overall, 133 (12%, 95% CI: 10.2–14.1) participants were HIV-positive, of whom 11 tested recent by MAA (BED-CEIA OD-n < 0.8 + LAg avidity OD-n < 1.5 + VL > 1000 copies/mL), giving an incidence of 1.46% (95% CI: 0.58–2.35) per year. This MAA-based incidence was similar to longitudinal KICoS1 incidence. Correlates of testing recent included sexually transmitted infection (STI) treatment history (OR = 3.94, 95% CI: 1.03–15.07) and syphilis seropositivity (OR = 10.15, 95% CI: 1.51–68.22). Overall, HIV-1 subtype A (63%), D (15%), C (3%), G (1%) and recombinants (18%), two monophyletic dyads and intrinsic viral mutations (V81I, V81I/V, V108I/V and K101Q) were observed. Viral diversity mirrored known patterns in this region, while resistance mutations reflected likely non-exposure to antiretroviral drugs. Management of STIs may help address ongoing HIV transmission in this region.

Newly diagnosed HIV-1 cases: Decreased proportion of recent infections based on a multi-assay algorithm combining two serological assays

Newly diagnosed HIV-1 cases: Decreased proportion of recent infections based on a multi-assay algorithm combining two serological assays

Community viral load, antiretroviral therapy coverage, and HIV incidence in India: a cross-sectional, comparative study

HIV incidence is the best measure of treatment-programme effectiveness, but its measurement is difficult and expensive. The concept of community viral load as a modifiable driver of new HIV infections has attracted substantial attention. We set out to compare several measures of community viral load and antiretroviral therapy (ART) coverage as correlates of HIV incidence in high-risk populations. We analysed data from a sample of people who inject drugs and men who have sex with men, who were participants of the baseline assessment of a cluster-randomised trial in progress across 22 cities in India (ClinicalTrials.gov number NCT01686750). We recruited the study population by use of respondent-driven sampling and did the baseline assessment at 27 community-based sites (12 for men who have sex with men and 15 for people who inject drugs). We estimated HIV incidence with a multiassay algorithm and calculated five community-based measures of HIV control: mean log10 HIV RNA in participants with HIV in a community either engaged in care (in-care viral load), aware of their status but not necessarily in care (aware viral load), or all HIV-positive individuals whether they were aware, in care, or not (population viral load); participants with HIV in a community with HIV RNA more than 150 copies per mL (prevalence of viraemia); and the proportion of participants with HIV who self-reported ART use in the previous 30 days (population ART coverage). All participants were tested for HIV, with additional testing in HIV-positive individuals. We assessed correlations between the measures and HIV incidence with Spearman correlation coefficients and linear regression analysis. Between Oct 1, 2012, and Dec 19, 2013, we recruited 26,503 participants, 12,022 men who have sex with men and 14,481 people who inject drugs. Median incidence of HIV was 0·87% (IQR 0·40-1·17) in men who have sex with men and 1·43% (0·60-4·00) in people who inject drugs. Prevalence of viraemia was more strongly correlated with HIV incidence (correlation 0·81, 95% CI 0·62-0·91; p<0·0001) than all other measures, although correlation was significant with aware viral load (0·59, 0·27-0·79; p=0·001), population viral load (0·51, 0·16-0·74; p=0·007), and population ART coverage (-0·54, -0·76 to -0·20; p=0·004). In-care viral load was not correlated with HIV incidence (0·29, -0·10 to 0·60; p=0·14). With regression analysis, we estimated that to reduce HIV incidence by 1 percentage point in a community, prevalence of viraemia would need to be reduced by 4·34%, and ART use in HIV-positive individuals would need to increase by 19·5%. Prevalence of viraemia had the strongest correlation with HIV incidence in this sample and might be a useful measure of the effectiveness of a treatment programme. US National Institutes of Health, Elton John AIDS Foundation.

Intensifying Antiretroviral Therapy With Raltegravir and Maraviroc During Early Human Immunodeficiency Virus (HIV) Infection Does Not Accelerate HIV Reservoir Reduction.

Background. Persistent human immunodeficiency virus (HIV) within the CD4+ T-cell reservoir is an obstacle to eradication. We hypothesized that adding raltegravir and maraviroc to standard combination antiretroviral therapy (cART) during early HIV infection could substantially reduce viral reservoirs as a step towards eradication.Methods. A prospective, randomized, double-blinded, placebo-controlled pilot trial enrolled 32 participants with documented early (<6 months) HIV infection to either standard cART (emtricitabine/tenofovir/lopinavir/ritonavir) or intensive cART (standard regimen + raltegravir/maraviroc). Human immunodeficiency virus reservoirs were assessed at baseline and at 48 weeks by (1) proviral DNA, (2) cell-associated RNA, and (3) replication-competent virus, all from purified blood CD4+ T cells, and (4) gut proviral DNA. A multiassay algorithm (MAA) on baseline sera estimated timing of infection.Results. Thirty individuals completed the study to the 48-week endpoint. The reduction in blood proviral burden was −1.03 log DNA copies/106 CD4+ T cells versus −.84 log in the standard and intensive groups, respectively (P = .056). Overall, there was no significant difference in the rate of decline of HIV-associated RNA, replication-competent virus in blood CD4+ T cells, nor proviral gut HIV DNA to 48 weeks. Individuals who presented with more recent HIV infection had significantly lower virus reservoirs, and cART tended to reduce their reservoirs to a greater extent.Conclusions. Intensive cART led to no additional reduction in the blood virus reservoir at 48 weeks compared with standard cART. Human immunodeficiency virus reservoir size is smaller earlier in HIV infection. Other novel treatment strategies in combination with early cART will be needed to eliminate the HIV latent reservoir.

High HIV prevalence and incidence among MSM across 12 cities in India.

To characterize prevalence, incidence, and associated correlates of HIV infection among MSM in 12 cities across India. Cross-sectional sample using respondent-driven sampling from September 2012 to June 2013. A total 12022 MSM (~1000/city) were recruited. Participants had to be at least 18 years, self-identify as male, and report oral/anal intercourse with a man in the prior year. HIV infection was diagnosed using three rapid tests. Cross-sectional HIV incidence was estimated using a multiassay algorithm. All estimates incorporate respondent-driven sampling-II weights. Median age was 25 years, 45% self-identified as 'panthi' (predominantly penetrative anal intercourse) and 30.6% reported being married to a woman. Weighted HIV prevalence was 7.0% (range: 1.7-13.1%). In multivariate analysis, significantly higher odds of HIV infection was observed among those who were older, had lower educational attainment, were practicing purely receptive anal sex or both receptive and penetrative sex, and those who were herpes simplex virus-2 positive. Of 1147 MSM who tested HIV positive, 53 were identified as recent HIV infections (annualized incidence = 0.87%; range = 0-2.2%). In multivariate analysis, injecting drugs in the prior 6 months, syphilis, and higher number of male partners and fewer female partners were significantly associated with recent HIV infection. We observed a high burden of HIV among MSM in India with tremendous diversity in prevalence, incidence, and risk behaviors. In particular, we observed high incidence in areas with relatively low prevalence suggesting emerging epidemics in areas not previously recognized to have high HIV burden.

High HIV burden among people who inject drugs in 15 Indian cities.

Injecting drug use has historically been the principal driver of the HIV epidemic in the northeast states of India. However, recent data indicate growing numbers of people who inject drugs (PWIDs) in north and central Indian cities. We conducted face-to-face surveys among PWIDs in seven northeast and eight north/central Indian cities using respondent-driven sampling. We used a rapid HIV-testing protocol to identify seropositive individuals and multiassay algorithm to identify those with recent infection. We used multilevel regression models that incorporated sampling weights and had random intercepts for site to assess risk factors for prevalent and incident (recent) HIV infection. We surveyed 14 481 PWIDs from 15 Indian cities between January and December 2013. Participants reported high rates of needle/syringe sharing. The median (site range) estimated HIV prevalence and incidence were 18.1% (5.9, 44.9) and 2.9 per 100 person-years (0, 12.4), respectively. HIV prevalence was higher in northeast sites, whereas HIV incidence was higher in north/central sites. The odds of prevalent HIV were over three-fold higher in women than in men. Other factors associated with HIV prevalence or incidence included duration since first injection, injection of pharmaceutical drugs, and needle/syringe sharing. The burden of HIV infection is high among PWIDs in India, and may be increasing in cities where injecting drug use is emerging. Women who inject drugs were at substantially higher risk for HIV than men - a situation that may be mediated by dual injection-related and sexual risks.

How can we better identify early HIV infections?

Detection of early HIV infections (EHIs), including acute HIV infection (AHI), is important for individual health, prevention of HIV transmission, and measurement of HIV incidence. We describe markers of EHI, diagnostic strategies for detecting these markers, and ways to incorporate these strategies into diagnostic and HIV incidence algorithms. For individual diagnosis in the USA and Europe, laboratory-based diagnostic algorithms increasingly incorporate fourth-generation HIV antigen tests, allowing for earlier detection. In some sub-Saharan African settings, symptom-based screening is being explored to identify subsets of persons at high risk for AHI. Point-of-care diagnostics designed for AHI detection are in the pipeline and, if validated, represent an opportunity for real-time AHI diagnosis. At the population level, multiassay algorithms are promising new strategies for estimating HIV incidence on the basis of several assays applied to cross-sectional samples. These algorithms can be developed to optimize performance, in addition to cost and logistical considerations. There are important recent advances in detection of EHIs at the individual and population levels. Applying optimal combinations of tests in diagnostic and HIV incidence algorithms is urgently needed to support the multiple goals derived from enhanced detection and discrimination of EHIs.

A comparison of two measures of HIV diversity in multi-assay algorithms for HIV incidence estimation.

BackgroundMulti-assay algorithms (MAAs) can be used to estimate HIV incidence in cross-sectional surveys. We compared the performance of two MAAs that use HIV diversity as one of four biomarkers for analysis of HIV incidence.MethodsBoth MAAs included two serologic assays (LAg-Avidity assay and BioRad-Avidity assay), HIV viral load, and an HIV diversity assay. HIV diversity was quantified using either a high resolution melting (HRM) diversity assay that does not require HIV sequencing (HRM score for a 239 base pair env region) or sequence ambiguity (the percentage of ambiguous bases in a 1,302 base pair pol region). Samples were classified as MAA positive (likely from individuals with recent HIV infection) if they met the criteria for all of the assays in the MAA. The following performance characteristics were assessed: (1) the proportion of samples classified as MAA positive as a function of duration of infection, (2) the mean window period, (3) the shadow (the time period before sample collection that is being assessed by the MAA), and (4) the accuracy of cross-sectional incidence estimates for three cohort studies.ResultsThe proportion of samples classified as MAA positive as a function of duration of infection was nearly identical for the two MAAs. The mean window period was 141 days for the HRM-based MAA and 131 days for the sequence ambiguity-based MAA. The shadows for both MAAs were <1 year. Both MAAs provided cross-sectional HIV incidence estimates that were very similar to longitudinal incidence estimates based on HIV seroconversion.ConclusionsMAAs that include the LAg-Avidity assay, the BioRad-Avidity assay, HIV viral load, and HIV diversity can provide accurate HIV incidence estimates. Sequence ambiguity measures obtained using a commercially-available HIV genotyping system can be used as an alternative to HRM scores in MAAs for cross-sectional HIV incidence estimation.

Effect of community-based voluntary counselling and testing on HIV incidence and social and behavioural outcomes (NIMH Project Accept; HPTN 043): a cluster-randomised trial.

Although several interventions have shown reduced HIV incidence in clinical trials, the community-level effect of effective interventions on the epidemic when scaled up is unknown. We investigated whether a multicomponent, multilevel social and behavioural prevention strategy could reduce HIV incidence, increase HIV testing, reduce HIV risk behaviour, and change social and behavioural norms. For this phase 3 cluster-randomised controlled trial, 34 communities in four sites in Africa and 14 communities in Thailand were randomly allocated in matched pairs to receive 36 months of community-based voluntary counselling and testing for HIV (intervention group) or standard counselling and testing alone (control group) between January, 2001, and December, 2011. The intervention was designed to make testing more accessible in communities, engage communities through outreach, and provide support services after testing. Randomisation was done by a computer-generated code and was not masked. Data were collected at baseline (n=14 567) and after intervention (n=56.683) by cross-sectional random surveys of community residents aged 18-32 years. The primary outcome was HIV incidence and was estimated with a cross-sectional multi-assay algorithm and antiretroviral drug screening assay. Thailand was excluded from incidence analyses because of low HIV prevalence. This trial is registered at ClinicalTrials.gov, number NCT00203749. The estimated incidence of HIV in the intervention group was 1.52% versus 1.81% in the control group with an estimated reduction in HIV incidence of 13.9% (relative risk [RR] 0.86, 95% CI 0.73-1.02; p=0.082). HIV incidence was significantly reduced in women older than 24 years (RR=0.70, 0.54-0.90; p=0.0085), but not in other age or sex subgroups. Community-based voluntary counselling and testing increased testing rates by 25% overall (12-39; p=0.0003), by 45% (25-69; p<0·0001) in men and 15% (3-28; p=0.013) in women. No overall effect on sexual risk behaviour was recorded. Social norms regarding HIV testing were improved by 6% (95% CI 3-9) in communities in the intervention group. These results are sufficiently robust, especially when taking into consideration the combined results of modest reductions in HIV incidence combined with increases in HIV testing and reductions in HIV risk behaviour, to recommend the Project Accept approach as an integral part of all interventions (including treatment as prevention) to reduce HIV transmission at the community level. US National Institute of Mental Health, the Division of AIDS of the US National Institute of Allergy and Infectious Diseases, and the Office of AIDS Research of the US National Institutes of Health.

Failure to Identify HIV-Infected Individuals in a Clinical Trial Using a Single HIV Rapid Test for Screening

Background: In the HIV Prevention Trials Network (HPTN) 061 study, 8 (2.3%) of 348 HIV-infected participants identified as HIV uninfected at study enrollment using a single HIV rapid test for screening were found to be HIV infected after additional testing. Objectives: To evaluate the performance of different HIV assays for detection of HIV infection in HPTN 061 participants with missed infection and individuals with viral suppression. Methods: Plasma samples from 8 HPTN 061 participants, 17 elite controllers, and 101 individuals on antiretroviral treatment (ART) were tested for HIV with 3 rapid tests, 2 laboratory-based immunoassays, and a Western blot assay. The HPTN 061 samples were also tested with 2 HIV RNA assays and an antiretroviral drug assay. Results: Of the 8 HPTN 061 participants with missed infection, 1 was an elite controller, 1 was taking ART, 2 were missed because of testing or clerical errors, 1 had recent HIV infection (identified using a multi-assay algorithm), and 3 had acute HIV infection. Two (1.7%) of 118 individuals with viral suppression (both taking ART) had at least 1 false-negative test. Conclusions: In clinical trials, HIV infections can be missed for a variety of reasons. Using more than one assay to screen for HIV infection may reduce the number of missed infections.

Performance of a Limiting-Antigen Avidity Enzyme Immunoassay for Cross-Sectional Estimation of HIV Incidence in the United States

BackgroundA limiting antigen avidity enzyme immunoassay (HIV-1 LAg-Avidity assay) was recently developed for cross-sectional HIV incidence estimation. We evaluated the performance of the LAg-Avidity assay alone and in multi-assay algorithms (MAAs) that included other biomarkers.Methods and FindingsPerformance of testing algorithms was evaluated using 2,282 samples from individuals in the United States collected 1 month to >8 years after HIV seroconversion. The capacity of selected testing algorithms to accurately estimate incidence was evaluated in three longitudinal cohorts. When used in a single-assay format, the LAg-Avidity assay classified some individuals infected >5 years as assay positive and failed to provide reliable incidence estimates in cohorts that included individuals with long-term infections. We evaluated >500,000 testing algorithms, that included the LAg-Avidity assay alone and MAAs with other biomarkers (BED capture immunoassay [BED-CEIA], BioRad-Avidity assay, HIV viral load, CD4 cell count), varying the assays and assay cutoffs. We identified an optimized 2-assay MAA that included the LAg-Avidity and BioRad-Avidity assays, and an optimized 4-assay MAA that included those assays, as well as HIV viral load and CD4 cell count. The two optimized MAAs classified all 845 samples from individuals infected >5 years as MAA negative and estimated incidence within a year of sample collection. These two MAAs produced incidence estimates that were consistent with those from longitudinal follow-up of cohorts. A comparison of the laboratory assay costs of the MAAs was also performed, and we found that the costs associated with the optimal two assay MAA were substantially less than with the four assay MAA.ConclusionsThe LAg-Avidity assay did not perform well in a single-assay format, regardless of the assay cutoff. MAAs that include the LAg-Avidity and BioRad-Avidity assays, with or without viral load and CD4 cell count, provide accurate incidence estimates.