Evaluation of the Abbott ARCHITECT HIV Ag/Ab combo assay for determining recent HIV-1 infection.

BACKGROUND: Given the challenges and costs associated with implementing HIV-1 incidence assay testing, there is great interest in evaluating the use of commercial HIV diagnostic tests for determining recent HIV infection. A diagnostic test with the capability of providing reliable data for the determination of recent HIV infection without substantial modifications to the test protocol would have a significant impact on HIV surveillance. The Abbott ARCHITECT HIV Ag/Ab Combo Assay is an antigen/antibody immunoassay, which meets the criteria as the first screening test in the recommended HIV laboratory diagnostic algorithm for the United States. METHODS: In this study, we evaluated the performance characteristics of the ARCHITECT HIV Ag/Ab Combo signal-to-cutoff ratio (S/Co) for determining recent infection, including estimation of the mean duration of recent infection (MDRI) and false recent rate (FRR), and selection of recency cutoffs. RESULTS: The MDRI estimates for the S/Co recency cutoff of 400 is within the 4 to 12 months range recommended for HIV incidence assays, and the FRR rate for this cutoff was 1.5%. Additionally, ARCHITECT Combo S/Co values were compared relative to diagnostic test results from two prior prospective HIV-1 diagnostic studies in order to validate the use of the S/Co for both diagnostic and recency determination. CONCLUSION: Dual-use of the ARCHITECT Combo assay data for diagnostic and incidence purposes would reduce the need for separate HIV incidence testing and allow for monitoring of recent infection for incidence estimation and other public health applications.

Altered Antibody Responses in Persons Infected with HIV-1 While Using Preexposure Prophylaxis.

Preexposure prophylaxis (PrEP) is an effective HIV prevention tool, although effectiveness is dependent upon adherence. It is important to characterize the impact of PrEP on HIV antibody responses in people who experience breakthrough infections to understand the potential impact on timely diagnosis and treatment. Longitudinal HIV-1-specific antibody responses were evaluated in 42 people who inject drugs (PWID) from the Bangkok Tenofovir Study (BTS) (placebo = 28; PrEP = 14) who acquired HIV while receiving PrEP. HIV-1 antibody levels and avidity to three envelope proteins (gp41, gp160, and gp120) were measured in the plasma using a customized Bio-Plex (Bio-Rad Laboratories, Hercules, CA) assay. A time-to-event analysis was performed for each biomarker to compare the distribution of times at which study subjects exceeded the recent/long-term assay threshold, comparing PrEP and placebo treatment groups. We fit mixed-effects models to identify longitudinal differences in antibody levels and avidity between groups. Overall, longitudinal antibody levels and avidity were notably lower in the PrEP breakthrough group compared to the placebo group. Time-to-event analyses demonstrated a difference in time to antibody reactivity between treatment groups for all Bio-Plex biomarkers. Longitudinal gp120 antibody levels within the PrEP breakthrough group were decreased compared to the placebo group. When accounting for PrEP adherence, both gp120 and gp160 antibody levels were lower in the PrEP breakthrough group compared to the placebo group. We demonstrate hindered envelope antibody maturation in PWID who became infected while receiving PrEP in the BTS, which has significant implications for HIV diagnosis. Delayed maturation of the antibody response to HIV may increase the time to detection for antibody-based tests. Clinical Trial Registration Number, NCT00119106.

A Multiplex HIV Incidence Assay for Inferring Recent HIV-1 Transmission and Time of Infection.

BACKGROUND: Laboratory assays for determining recent HIV-1 infection are an important public health tool for aiding in the estimation of HIV incidence. Some incidence assay analytes are remarkably predictive of time since seroconversion and may be useful for additional applications, such as predicting recent transmission events during HIV outbreaks and informing prevention strategies. METHODS: Plasma samples (n = 154) from a recent HIV-1 outbreak in a rural community in Indiana were tested with the customized HIV-1 Multiplex assay, based on the Bio-Rad Bio-Plex platform, which measures antibody response to HIV envelope antigens, gp120, gp160, and gp41. Assay cutoffs for each analyte were established to determine whether an individual seroconverted within 30, 60, or 90 days of the sample collection date. In addition, a novel bioinformatics method was implemented to infer infection dates of persons newly diagnosed with HIV during the outbreak. RESULTS: Sensitivity/specificity of the HIV-1 Multiplex assay for predicting seroconversion within 30, 60, and 90 days, based on a training data set, was 90.5%/95.4%, 94.1%/90%, and 89.4%/82.9%, respectively. Of 154 new diagnoses in Indiana between December 2014 and August 2016, the majority (71%) of recent infections (≤3 months since seroconversion) were identified between February and May 2016. The epidemiologic curve derived from the bioinformatics analysis indicated HIV transmission began as early as 2010, grew exponentially in 2014, and leveled off in April 2015. CONCLUSIONS: The HIV-1 Multiplex assay has the potential to identify and monitor trends in recent infection during an epidemic to assess the efficacy of programmatic or treatment interventions.

Characterization of real-time microarrays for simultaneous detection of HIV-1, HIV-2, and hepatitis viruses.

Real-time PCR assays for nucleic acid testing (NAT) of hepatitis viruses A-E and for HIV-1 and HIV-2 have been developed; however, a multiplex assay that can simultaneously detect all of these agents is not yet available. Standardized TaqMan assays for detection of hepatitis viruses A-E have been described and applied to TaqMan Array Cards (TAC) which are capable of multiple pathogen detection using a single set of optimized PCR conditions. Assays for three gene regions of HIV-1 (long-terminal repeat (LTR), gag, and polymerase) and HIV-2 (overlap of LTR and gag, protease and integrase) were designed using the hepatitis assay conditions. Nucleic acid extracts of HIV-1-infected samples (44 plasma, 41 whole blood, 20 HIV-1 viral stocks) were tested on the TAC cards; 98 were reactive (92%) with 70 in multiple gene regions. Twenty-four of the 27 (89%) HIV-2 specimens (10 plasma, 1 PBMC lysate, 6 whole blood and 10 plasmids containing HIV-2 polymerase) were detected on TAC. No HIV or hepatitis virus sequences were detected in 30 HIV-negative samples (specificity 100%). Three HBV and 18 HCV co-infections were identified in the HIV-1-infected specimens. Multi-pathogen detection using TAC could provide a rapid, sensitive and more efficient method of surveying for a variety of infectious disease nucleic acids.

A multiplexed RT-LAMP assay for detection of group M HIV-1 in plasma or whole blood.

Isothermal nucleic acid amplification techniques, such as reverse-transcription loop-mediated isothermal amplification (RT-LAMP), exhibit characteristics that are suitable for the development of a rapid, low-cost NAT that can be used at the POC. For demonstration of utility for global use, studies are needed to validate the performance of RT-LAMP for the detection of divergent subtypes. In this study, we designed and evaluated multiplexed HIV-1 integrase RT-LAMP primers to detect subtypes within group M, along with an RNase P positive internal processing and amplification control. Using a panel of 26 viral isolates representing the major circulating subtypes, we demonstrated detection of all isolates of subtypes A1, C, D, F1, F2, G, CRF01_AE, CRF02_AG, and two unique recombinant forms (URFs). A whole blood panel created with one representative isolate of each subtype was successfully amplified with the group M HIV-1 integrase and RNase P internal control primers. The group M HIV-1 RT-LAMP assay was further evaluated on 61 plasma specimens obtained from persons from Cameroon and Uganda. The sequence-conserved group M HIV-1 RT-LAMP primers, coupled to a low-cost amplification device, may improve diagnosis of acute infection at the POC and provide timely confirmation of HIV status.

A multiplex assay for detection of SHIV plasma and mucosal IgG and IgA.

Evaluating antibody maturation provides valuable data to characterize immune responses to HIV infection and can provide insight into biomedical intervention efficacy. It is important to develop assays that evaluate antibody maturation in both plasma and mucosal compartments. The nonhuman primate model provides a controlled system to collect temporal data that are integral to assessing intervention strategies. We report the development of a novel multiplex assay, based on the Bio-Plex platform, to evaluate plasma and mucosal IgG and IgA avidity and maturation against simian/human immunodeficiency virus (SHIV) in this controlled system. Vaginal mucosa and plasma samples were collected from a prior study evaluating the efficacy of a tenofovir disoproxil fumarate (TDF) intravaginal ring (IVR) against SHIVSF162P3 challenge in female pigtailed macaques. For validation of the multiplex assay, specimens from six SHIV-infected placebo animals and one TDF breakthrough animal were evaluated. For SHIV and HIV envelope analytes, antibody levels and avidity in both compartments continued to mature post-infection. Maturation of IgG and IgA levels was similar in each compartment, however, mucosal antibody levels tended to be more variable. This SHIV assay elucidates IgG/IgA antibody kinetics in the plasma and vaginal mucosa and will be a valuable tool in vaccine and other biomedical intervention studies in the nonhuman primate model.

Performance characteristics of an antibody-based multiplex kit for determining recent HIV-1 infection.

The availability of reliable laboratory methods for determining recent HIV infection is vital for accurate estimation of population-based incidence. The mean duration of recent infection (MDRI) and false recent rate (FRR) are critical parameters for HIV incidence assays, as they impact HIV incidence estimates and provide a measure of assay performance. The HIV-1 Multiplex assay is an in-house developed, magnetic bead-based assay that measures virus-specific antibody levels and avidity to multiple analytes. To ensure quality control and to facilitate transfer of the assay to external laboratories or testing facilities, the in-house assay has been adapted and produced in kit form. Here, we describe the performance characteristics of the multiplex kit and demonstrate the stability of the kit components over a one-year period. Two statistical methods were employed to estimate the MDRI of the individual analytes and five different algorithms, combining multiple analyte values. The MDRI estimates for the individual analytes and five algorithms were all between 200 and 300 days post-seroconversion, with no notable difference between the two statistical approaches. All five algorithms exhibited a 0% FRR with specimens from long-term, subtype B HIV-1-infected individuals. The assay parameters described in this study provide the necessary tools to implement the HIV-1 multiplex assay and improves the utility of the assay for field use.

Single-use, electricity-free amplification device for detection of HIV-1.

Early and accurate diagnosis of HIV is key for the reduction of transmission and initiation of patient care. The availability of a rapid nucleic acid test (NAT) for use at the point-of-care (POC) will fill a gap in HIV diagnostics, improving the diagnosis of acute infection and HIV in infants born to infected mothers. In this study, we evaluated the performance of non-instrumented nucleic acid amplification, single-use disposable (NINA-SUD) devices for the detection of HIV-1 in whole blood using reverse-transcription, loop-mediated isothermal amplification (RT-LAMP) with lyophilized reagents. The NINA-SUD heating device harnesses the heat from an exothermic chemical reaction initiated by the addition of saline to magnesium iron powder. Reproducibility was demonstrated between NINA-SUD units and comparable, if not superior, performance for detecting clinical specimens was observed as compared to the thermal cycler. The stability of the lyophilized HIV-1 RT-LAMP reagents was also demonstrated following storage at -20, 4, 25, and 30°C for up to one month. The single-use, disposable NAT minimizes hands-on time and has the potential to facilitate HIV-1 testing in resource-limited settings or at the POC.

Short Communication: Persistence of HIV Antibody Avidity in the Presence of Antiretroviral Therapy.

The effects of antiretroviral therapy (ART) on the performance of HIV incidence assays have been well documented. To improve upon current assay approaches or focus the development of future assays, studies are needed to characterize the effects of ART on all candidate HIV incidence assays. In this study, we compared the performance of three antibody avidity-based HIV incidence assays, the Limiting Antigen (LAg), Bio-Rad Avidity, and HIV-1 Multiplex assays, using a well-defined cohort of recent HIV-1 seroconverters composed of ART-naive HIV-1-infected individuals and those who received ART early or delayed in the course of infection. Differences in the performance of all three avidity-based incidence assays were noted with study subjects who received ART. The LAg assay and Multiplex total antibody measurements (nMFI) exhibited similar kinetics in reactivity, as these assays tended to fluctuate with changes in viral load. In the early ART group, all seven subjects remained recent by both assays at time points >1 year postseroconversion, and assay values declined dramatically postdelayed ART initiation. In contrast, the two-well, antibody-dissociation avidity assays, Bio-Rad Avidity and Multiplex avidity index (AI) measurements, continued to mature in the early ART group, although blunted relative to the ART-naive group, and assay values remained stable after delayed ART initiation. In summary, although the HIV incidence assays evaluated in this study are all designed to measure antibody avidity, each assay is affected differently by ART-induced virus suppression, presumably because of the distinct assay formats and procedures for measuring avidity.

Detection of Acute HIV-1 Infection by RT-LAMP.

A rapid, cost-effective diagnostic test for the detection of acute HIV-1 infection is highly desired. Isothermal amplification techniques, such as reverse-transcription loop-mediated isothermal amplification (RT-LAMP), exhibit characteristics that are ideal for the development of a rapid nucleic acid amplification test (NAAT) because they are quick, easy to perform and do not require complex, dedicated equipment and laboratory space. In this study, we assessed the ability of the HIV-1 RT-LAMP assay to detect acute HIV infection as compared to a representative rapid antibody test and several FDA-approved laboratory-based assays. The HIV-1 RT-LAMP assay detected seroconverting individuals one to three weeks earlier than a rapid HIV antibody test and up to two weeks earlier than a lab-based antigen/antibody (Ag/Ab) combo enzyme immunoassay (EIA). RT-LAMP was not as sensitive as a lab-based qualitative RNA assay, which could be attributed to the significantly smaller nucleic acid input volume. To our knowledge, this is the first demonstration of detecting acute HIV infection using the RT-LAMP assay. The availability of a rapid NAAT, such as the HIV-1 RT-LAMP assay, at the point of care (POC) or in laboratories that do not have access to large platform NAAT could increase the percentage of individuals who receive an acute HIV infection status or confirmation of their HIV status, while immediately linking them to counseling and medical care. In addition, early knowledge of HIV status could lead to reduced high-risk behavior at a time when individuals are at a higher risk for transmitting the virus.

Longitudinal analysis of HIV-1-specific antibody responses.

Laboratory assays for determining recent HIV-1 infection are of great public health importance for aiding in the estimation of HIV incidence. Concerns have been raised about the potential for misclassification with serology-based assays due to fluctuations in the antibody response, particularly following progression to AIDS. We characterized longitudinal antibody responses to HIV using a cohort of men who have sex with men (MSM) sampled for up to 17 years, in which 57% of the 65 study subjects included in the current analyses progressed to AIDS during the study period. Envelope-specific total IgG antibody levels, avidity, and p24-specific IgG3 levels were evaluated using a multiplexed Bio-Plex assay. For the majority of the analytes, no significant difference in IgG reactivity was observed between AIDS and non-AIDS specimens. Although a slight decline in gp120 reactivity was noted with decreasing CD4(+) T cell count, the drop in assay values was relatively minimal and would likely not lead to an increase in the misclassification rate of the assay. A peak in HIV-1 p24 IgG3 levels was observed during early infection, as confirmed by testing 1,216 specimens from 342 recent seroconverters with the Bio-Plex assay. As expected, IgG3 reactivity declined with disease progression and decreasing CD4(+) T cell count in the MSM cohort; however, 37% of the study subjects exhibited relatively high IgG3 levels late in the course of infection.

Evaluation of dried blood spots with a multiplex assay for measuring recent HIV-1 infection.

Laboratory-based HIV tests for recent infection (TRIs), which primarily measure a specific serological biomarker(s) that distinguishes recent from long-term HIV infection, have facilitated the estimation of population-based incidence. Dried blood spots (DBS) on filter paper are an attractive sample source for HIV surveillance, given the simplified and cost-effective methods of specimen collection, storage, and shipment. Here, we evaluated the use of DBS in conjunction with an in-house multiplex TRI, the HIV-1-specific Bio-Plex assay, which measures direct antibody binding and avidity to multiple HIV-1 analytes. The assay performance was comparable between matched plasma and DBS samples from HIV-1 infected individuals obtained from diverse sources. The coefficients of variation, comparing the median antibody reactivity for each analyte between plasma and DBS, ranged from 2.78% to 9.40% and the correlation coefficients between the two sample types ranged from 0.89 to 0.97, depending on the analyte. The correlation in antibody reactivity between laboratory and site-prepared DBS for each analyte ranged from 0.87 to 0.98 and from 0.90 to 0.97 between site-prepared DBS and plasma. The correlation in assay measures between plasma and DBS indicate that the sample types can be used interchangeably with the Bio-Plex format, without negatively impacting the misclassification rate of the assay.

Reference panel of cloned HIV-2 plasmid DNA for nucleic acid assay development, evaluation, and quality monitoring.

BACKGROUND: Currently, no FDA-approved HIV-2 nucleic acid assay is commercially available in the United States, although several laboratories have developed in-house assays to confirm HIV-2 infections. A major limitation in the development of novel HIV-2 diagnostic assays is the lack of reference materials that can be used to evaluate, optimize, and monitor assay performance. STUDY DESIGN: Eleven viral stocks of HIV-2 isolates from various West African countries, including the Ivory Coast, Senegal, and Guinea-Bissau, were used to clone the entire LTR and pol regions from each virus. RESULTS: We successfully cloned, sequenced, and group classified 22 HIV-2 DNA plasmids including 11 full length LTR (∼849 bp) and 11 pol (∼2995 bp) sequences. There were eight HIV-2 group A and three group B in both the LTR and pol regions. CONCLUSIONS: This reference panel provides a robust, quantifiable, renewable, and non-infectious set of reagents that can be used for the development and evaluation of new HIV-2 molecular diagnostic assays and quality assurance and quality control reagents for use in the clinical laboratories.

Real-Time Detection of HIV-2 by Reverse Transcription-Loop-Mediated Isothermal Amplification.

Currently, there are no FDA-approved nucleic acid amplification tests (NAATs) for the detection or confirmation of HIV-2 infection. Here, we describe the development of a real-time assay for the detection of HIV-2 DNA and RNA using reverse transcription-loop-mediated isothermal amplification (RT-LAMP) and the ESEQuant tube scanner, a portable isothermal amplification/detection device.

Inter-laboratory assessment of a prototype multiplex kit for determination of recent HIV-1 infection.

BACKGROUND: Accurate and reliable laboratory-based assays are needed for estimating HIV-1 incidence from cross-sectional samples. We recently described the development of a customized, HIV-1-specific Bio-Plex assay that allows for the measurement of HIV-specific antibody levels and avidity to multiple analytes for improved HIV-1 incidence estimates. METHODS: To assess intra- and inter-laboratory assay performance, prototype multiplex kits were developed and evaluated by three distinct laboratories. Longitudinal seroconversion specimens were tested in parallel by each laboratory and kit performance was compared to that of an in-house assay. Additionally, the ability of the kit to distinguish recent from long-term HIV-1 infection, as compared to the in-house assay, was determined by comparing the reactivity of known recent (infected <6 months) and long-term (infected >12 months) drug naïve specimens. RESULTS: Although the range of reactivity for each analyte varied between the prototype kit and in-house assay, a measurable distinction in reactivity between recent and long-term specimens was observed with both assays in all three laboratories. Additionally, kit performance was consistent between all three laboratories. The intra-assay coefficient of variation (CV), between sample replicates for all laboratories, ranged from 0.5% to 6.1%. The inter-laboratory CVs ranged from 8.5% to 21.3% for gp160-avidity index (a) and gp120-normalized mean fluorescent intensity (MFI) value (n), respectively. CONCLUSION: We demonstrate the feasibility of producing a multiplex kit for measuring HIV antibody levels and avidity, with the potential for improved incidence estimates based on multi-analyte algorithms. The availability of a commercial kit will facilitate the transfer of technology among diverse laboratories for widespread assay use.

Evaluation of a multiplex assay for estimation of HIV-1 incidence.

OBJECTIVES: Accurate methods of estimating HIV-1 incidence are critical for monitoring the status of the epidemic and the impact of prevention strategies. Although several laboratory-based tests have been developed strictly for this purpose, several limitations exist and improved methods or technologies are needed. We sought to further optimize a previously described bead-based, HIV-1-specific multiplex assay with the capability of measuring multiple immune responses for determining recent infection. METHODS: We refined the customized HIV-1 Bio-Plex assay by determining cutoffs and mean durations of recency (MDR), based on the reactivity to longitudinal seroconversion specimens (n = 1347) from 311 ART-naïve, HIV-1-infected subjects. False-recent rates (FRRs) were calculated for various long-term cohorts, including AIDS patients, individuals on ART, and subtype C specimens. Incidence was estimated for each individual assay analyte from a simulated population with a known incidence of 1%. For improved incidence estimates, multi-analyte algorithms based on combinations of 3 to 6 analytes were evaluated and compared to the performance of each individual analyte. RESULTS: The MDR for the six analytes varied from 164.2 to 279.4 days, while the multi-analyte algorithm MDRs were less variable with a minimum and maximum value of 228.4 and 277.9 days, respectively. The FRRs for the 7 multi-analyte algorithms evaluated in this study varied from 0.3% to 3.1%, in a population of ART-naïve, long-term individuals. All algorithms yielded improved incidence estimates as compared to the individual analytes, predicting an incidence of 0.95% to 1.02%. CONCLUSIONS: The HIV-specific multiplex assay described here measures several distinct immune responses in a single assay, allowing for the consideration of multi-analyte algorithms for improved HIV incidence estimates.

Isothermal amplification using a chemical heating device for point-of-care detection of HIV-1.

BACKGROUND: To date, the use of traditional nucleic acid amplification tests (NAAT) for detection of HIV-1 DNA or RNA has been restricted to laboratory settings due to time, equipment, and technical expertise requirements. The availability of a rapid NAAT with applicability for resource-limited or point-of-care (POC) settings would fill a great need in HIV diagnostics, allowing for timely diagnosis or confirmation of infection status, as well as facilitating the diagnosis of acute infection, screening and evaluation of infants born to HIV-infected mothers. Isothermal amplification methods, such as reverse-transcription, loop-mediated isothermal amplification (RT-LAMP), exhibit characteristics that are ideal for POC settings, since they are typically quicker, easier to perform, and allow for integration into low-tech, portable heating devices. METHODOLOGY/SIGNIFICANT FINDINGS: In this study, we evaluated the HIV-1 RT-LAMP assay using portable, non-instrumented nucleic acid amplification (NINA) heating devices that generate heat from the exothermic reaction of calcium oxide and water. The NINA heating devices exhibited stable temperatures throughout the amplification reaction and consistent amplification results between three separate devices and a thermalcycler. The performance of the NINA heaters was validated using whole blood specimens from HIV-1 infected patients. CONCLUSION: The RT-LAMP isothermal amplification method used in conjunction with a chemical heating device provides a portable, rapid and robust NAAT platform that has the potential to facilitate HIV-1 testing in resource-limited settings and POC.

Development and characterization of a bead-based, multiplex assay for estimation of recent HIV type 1 infection.

Estimation of HIV-1 incidence is an important public health tool for understanding the status of the epidemic, identifying high-risk populations, and assessing various intervention strategies. Several laboratory-based methods have been developed for distinguishing recent from long-term HIV-1 infection; however, each exhibits some degree of misclassification, particularly among AIDS patients and those taking antiretroviral therapy (ART). To improve upon the limitations associated with measuring responses to a single analyte, we have developed a bead-based, multiplex assay for determination of HIV recent infection based on total antibody binding and antibody avidity to multiple analytes. An HIV-specific, multiplex panel was created by coupling the recombinant HIV-1 proteins p66, gp120, gp160, and gp41 to Bio-Plex COOH microspheres. Longitudinal plasma specimens from recent seroconverters were tested for reactivity to the coupled microspheres using the Bio-Plex 200 System. For each analyte, HIV-specific antibody binding and avidity increased for 1-2 years post-seroconversion, leading to a significant difference in reactivity between recent and long-term specimens. While the potential for misclassification of individuals diagnosed with AIDS or receiving ART appears to be minimal with avidity measures, the impact on total antibody binding was variable, depending on the individual analyte. This bead-based, HIV-specific multiplex assay measures several distinct immune responses in a single assay plate, allowing for sampling of multiple analytes in the determination of recent infection, which could aid in the development of improved statistical methods or algorithms that will more accurately estimate HIV incidence.

Delayed maturation of antibody avidity but not seroconversion in rhesus macaques infected with simian HIV during oral pre-exposure prophylaxis.

BACKGROUND: Pre-exposure prophylaxis (PrEP) is a novel intervention strategy for the prevention HIV transmission. Because several clinical trials are at various stages of completion, it is important to understand the impact of PrEP treatment on the development of the immune response to HIV, particularly in individuals who exhibit breakthrough infections despite PrEP. METHODS: A model of HIV infection, using rhesus macaques and the simian/human immunodeficiency virus (SHIV), was used to evaluate the effects of PrEP on the evolution of the humoral immune response. Time to seroconversion, neutralizing and binding antibody levels, and antibody avidity were measured in 12 rhesus macaques infected during daily or intermittent PrEP with FTC (emtricitabine) or Truvada (FTC/tenofovir combination) and compared with 11 untreated, simian HIV-infected controls. RESULTS: Macaques that became infected while receiving PrEP exhibited significantly lower peak virus loads during acute infection as compared with untreated animals. Although the timing of seroconversion and SHIV binding and neutralizing antibody levels were not impacted by treatment, lower maturation rates of antibody avidity for anti-p27, gp120, gp160, and gp41 were observed. CONCLUSIONS: This study suggests that reduced virus loads associated with PrEP treatment have little impact on timing of seroconversion and neutralizing/binding antibody levels; however, maturation of antibody avidity was suppressed.

Sequence-specific detection method for reverse transcription, loop-mediated isothermal amplification of HIV-1.

HIV diagnosis at the point-of-care or in resource-limited settings poses considerable challenges due to time and cost limitations. Currently, nucleic acid-based tests are the only reliable method for diagnosing recent infections during the window period post-infection and pre-seroconversion, but these tests are only suitable for well-equipped laboratory settings. The reverse transcription loop-mediated isothermal amplification (RT-LAMP) technology exhibits characteristics that are ideal for the development of a rapid, cost-effective nucleic acid-based test for detection of HIV DNA and RNA. In this study, a sequence-specific detection method was developed for immediate, naked-eye visualization of RT-LAMP products with high sensitivity and specificity. The rapid detection method was incorporated into the HIV-1-specific RT-LAMP assay and validated using minute volumes of whole blood from HIV-1-infected individuals. Together with the minimal sample preparation time and one-step, isothermal amplification reaction, the sequence-specific detection method adds to the overall versatility of the RT-LAMP assay and enhances the applicability for use at point-of-care or resource-limited sites.