The LAg-Avidity EIA differentiates between recent and long-term HIV infection. part of the Botswana Combination Prevention Project (BCPP), from October 2013 CNovember 2015. Data and samples from your baseline household survey were used to estimate cross-sectional HIV incidence, following an algorithm that combined Limiting-Antigen Avidity Assay (LAg-Avidity EIA), ART status (documented or by screening ARV drugs in plasma) and HIV-1 RNA weight. The LAg-Avidity EIA cut-off normalized optical density (ODn) was set at 1.5. The HIV-1 RNA cut-off was set at 400 copies/mL. For estimation purposes, the Mean Period of Recent Contamination was 130 days and the False Recent Rate (FRR) was Thbd evaluated at values of either 0 or 0.39%. Results Among 12,610 individuals participating in the baseline household survey, HIV status was available for 12,570 participants and 3,596 of them were HIV positive. LAg-Avidity EIA data was generated for 3,581 (99.6%) of HIV-positive participants. Of 326 participants with ODn 1.5, 278 individuals were receiving ART verified through paperwork and were considered to represent gamma-Mangostin longstanding HIV infections. Among the remaining 48 participants who reported no use of ART, 14 experienced an HIV-1 RNA weight 400 copies/mL gamma-Mangostin (including 3 participants with ARVs in plasma) and were excluded, as potential elite/viremic controllers or undisclosed ART. Thus, 34 LAg-Avidity-EIA-recent, ARV-na?ve individuals with detectable HIV-1 RNA ( 400 copies/mL) were classified as individuals with recent HIV infections. The annualized HIV incidence among 16C64 12 months aged adults was estimated at 1.06% (95% CI 0.68C1.45%) with zero FRR, and at 0.64% (95% CI 0.24C1.04%) using a previously defined FRR of 0.39%. Within a subset of more youthful individuals 16C49 years old, the annualized HIV incidence was estimated at 1.29% (95% CI 0.82C1.77%) with zero FRR, and at 0.90% (95% CI 0.42C1.38%) with FRR set to 0.39%. Conclusions Using a cross-sectional estimate of HIV incidence from 2013C2015, we found that at the time of near achievement of the UNAIDS 90-90-90 targets, ~1% of adults (age 16C64 years) in Botswanas rural and peri-urban communities became HIV infected annually. Introduction Botswana has been hard hit by the HIV-epidemic, with the third highest HIV prevalence worldwide among adults age 15C49, after Lesotho and Swaziland . Botswana appears to be approaching the UNAIDS 90-90-90 HIV screening, treatment, and viral suppression targets . These high levels of protection have led to significant reductions in HIV-related mortality [1, 3C5]. In June 2016 Botswana adopted the gamma-Mangostin World Health Organization (WHO) recommendation to provide Universal Test and Treat (UTT) . The success of UTT could be measured by reduction in HIV incidence [7C25]. Monitoring of HIV incidence is a critical tool for assessment and evaluation the impact of HIV prevention and treatment programs. Prospective longitudinal cohorts remain the gold standard for assessing HIV incidence. However, this approach is time consuming, costly and prone to selection and observational biases [8, 26]. Biomarkers of recent HIV infection that can be detected in cross-sectional samples represent a viable alternative to longitudinal cohort studies. Serological and molecular biomarkers could be combined in multi-assay algorithm (MAA). An optimized MAA with high sensitivity and specificity can discriminate between recent and established HIV infections in cross-sectional sample [27C29][8, 9, 25, 30C36]. Recent advances in design and development of MAA have facilitated estimating of HIV incidence in cross-sectional surveys with improved accuracy [7, 16, 22, 37]. In this study, we estimated HIV incidence using a baseline cross-sectional sample from your Botswana Combination Prevention Project (BCPP; the study) [2, 38, 39]. BCPP is an ongoing pair-matched, cluster-randomized clinical trial in 30 rural and peri-urban communities across Botswana. The primary question of the study is reduction in the cumulative HIV incidence as a result of combination prevention interventions that included enhanced HIV screening and counseling (HTC) campaigns, linkage to care, antiretroviral treatment (ART), strengthened male circumcision (MC) and enhanced prevention of gamma-Mangostin mother-to-child transmission.