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HIV-1 RNA genetic diversity predicts time since infection which is important for clinical care and research. It's unclear, however, whether proviral DNA genetic diversity sampled under suppressive antiretroviral therapy can be used for this purpose. We tested whether proviral genetic diversity from NGS sequences predicts time since infection and recency in 221 people with HIV-1 with known infection time. Proviral diversity was significantly associated with time since infection (p<5*10-07, R2 up to 25%) and predictive of treatment initiation during recent infection (AUC-ROC up to 0.85). This shows the utility of proviral genetic diversity as a proxy for time since infection.

BACKGROUND: Clinical and laboratory monitoring of patients on antiretroviral therapy is an integral part of HIV care and determines whether treatment needs enhanced adherence or modification of the drug regimen. However, different monitoring and treatment strategies carry different costs and health consequences. MATERIALS AND METHODS: The SIMPL’HIV study was a randomised trial that assessed the non-inferiority of dual maintenance therapy. The co-primary outcome was a comparison of costs over 48 weeks of dual therapy with standard antiretroviral therapy and the costs associated with a simplified HIV care approach (patient-centred monitoring [PCM]) versus standard, tri-monthly routine monitoring. Costs included outpatient medical consultations (HIV/non-HIV consultations), non-medical consultations, antiretroviral therapy, laboratory tests and hospitalisation costs. PCM participants had restricted immunological and blood safety monitoring at weeks 0 and 48, and they were offered the choice to complete their remaining study visits via a telephone call, have medications delivered to a specified address, and to have blood tests performed at a location of their choice. We analysed the costs of both strategies using invoices for medical consultations issued by the hospital where the patient was followed, as well as those obtained from health insurance companies. Secondary outcomes included differences between monitoring arms for renal function, lipids and glucose values, and weight over 48 weeks. Patient satisfaction with treatment and monitoring was also assessed using visual analogue scales. RESULTS: Of 93 participants randomised to dolutegravir plus emtricitabine and 94 individuals to combination antiretroviral therapy (median nadir CD4 count, 246 cells/mm3; median age, 48 years; female, 17%),patient-centred monitoring generated no substantial reductions or increases in total costs (US$ –421 per year [95% CI –2292 to 1451]; p = 0.658). However, dual therapy was significa

BACKGROUND T-cell responses during chronic viral infections become exhausted, which is reflected by upregulation of inhibitory receptors (iRs) and increased interleukin 10 (IL-10). We assessed 2 iRs-PD-1 (programmed cell death protein 1) and Tim-3 (T-cell immunoglobulin and mucin domain-containing protein 3)-and IL-10 mRNAs in peripheral blood mononuclear cells (PBMCs) and their soluble analogs (sPD-1, sTim-3, and IL-10) in plasma in chronic HIV-1/hepatitis C virus (HCV) coinfection and explored the effect of HCV treatment on these markers. We also aimed to establish whether iR expression may be determined by the HCV CD8$^{+}$ T-cell immunodominant epitope sequence. METHODS Plasma and PBMCs from 31 persons with chronic HIV-1/HCV coinfection from the Swiss HIV Cohort Study were collected before and after HCV treatment. As controls, 45 persons who were HIV-1 negative with chronic HCV infection were recruited. Exhaustion markers were assessed by enzyme-linked immunosorbent assay in plasma and by quantitative reverse transcription polymerase chain reaction in PBMCs. Analysis of an HCV epitope sequence was conducted by next-generation sequencing: HLA-A*02-restricted NS3$_{1073-1081}$ and NS3$_{1406-1415}$ and HLA-A*01-restricted NS3$_{1436-1444}$. RESULTS The study revealed higher plasma sPD-1 (P = .0235) and IL-10 (P = .002) levels and higher IL-10 mRNA in PBMCs (P = .0149) in HIV-1/HCV coinfection. A decrease in plasma sPD-1 (P = .0006), sTim-3 (P = .0136), and IL-10 (P = .0003) and Tim-3 mRNA in PBMCs (P = .0210) was observed following successful HCV treatment. Infection with the HLA-A*01-restricted NS3$_{1436-1444}$ ATDALMTGY prototype variant was related to higher sTim-3 levels than infection with the ATDALMTGF escape variant (P = .0326). CONCLUSIONS The results underscore the synergistic effect of coinfection on expression of exhaustion markers, their reduction following successful HCV treatment and imply that iR levels may operate on an epitope-specific manner.

BACKGROUND: Starting combination antiretroviral therapy (cART) during primary human immunodeficiency virus type 1 (HIV-1) infection results in a smaller HIV-1 latent reservoir, reduced immune activation, and less viral diversity compared to starting cART during chronic infection. We report results of a 4-year study designed to determine whether these properties would allow sustained virological suppression after simplification of cART to dolutegravir (DTG) monotherapy. METHODS: EARLY-SIMPLIFIED is a randomized, open-label, noninferiority trial. People with HIV (PWH) who started cART <180 days after a documented primary HIV-1 infection with suppressed viral load were randomized (2:1) to DTG monotherapy with 50 mg daily or continuation of cART. The primary endpoints were the proportion of PWH with viral failure at 48, 96, 144, and 192 weeks; noninferiority margin was 10%. After 96 weeks, randomization was lifted and patients were permitted to switch treatment groups as desired. RESULTS: Of 101 PWH randomized, 68 were assigned to DTG monotherapy and 33 to cART. At week 96 in the per-protocol population, 64/64 (100%) showed virological response in the DTG monotherapy group versus 30/30 (100%) in the cART group (difference, 0.00%; upper bound of 95% confidence interval 6.22%). This demonstrated noninferiority of DTG monotherapy at the prespecified level. At week 192, the study end, no virological failure occurred in either group during 13 308 and 4897 person weeks of follow-up for the DTG monotherapy (n = 80) and cART groups, respectively. CONCLUSIONS: This trial suggests that early cART initiation during primary HIV infection allows sustained virological suppression after switching to DTG monotherapy.

BACKGROUND: Genotypic resistance testing (GRT) is routinely performed upon diagnosis of HIV-1 infection or during virological failure using plasma viral RNA. An alternative source for GRT could be cellular HIV-1 DNA. OBJECTIVES: A substantial number of participants in the Swiss HIV Cohort Study (SHCS) never received GRT. We applied a method that enables access to the near full-length proviral HIV-1 genome without requiring detectable viraemia. METHODS: Nine hundred and sixty-two PBMC specimens were received. Our two-step nested PCR protocol was applied to generate two overlapping long-range amplicons of the HIV-1 genome, sequenced by next-generation sequencing (NGS) and analysed by MinVar, a pipeline to detect drug resistance mutations (DRMs). RESULTS: Six hundred and eighty-one (70.8%) of the samples were successfully amplified, sequenced and analysed by MinVar. Only partial information of the pol gene was contained in 82/681 (12%), probably due to naturally occurring deletions in the proviral sequence. All common HIV-1 subtypes were successfully sequenced. We detected at least one major DRM at high frequency (≥15%) in 331/599 (55.3%) individuals. Excluding APOBEC-signature (G-to-A mutation) DRMs, 145/599 (24.2%) individuals carried at least one major DRM. RT-inhibitor DRMs were most prevalent. The experienced time on ART was significantly longer in DRM carriers (P = 0.001) independent of inclusion or exclusion of APOBEC-signature DRMs. CONCLUSIONS: We successfully applied a reliable and efficient method to analyse near full-length HIV-1 proviral DNA and investigated DRMs in individuals with undetectable or low viraemia. Additionally, our data underscore the need for new computational tools to exclude APOBEC-related hypermutated NGS sequence reads for reporting DRMs.

While combination therapy completely suppresses HIV-1 replication in blood, functional virus persists in CD4$^{+}$ T cell subsets in non-peripheral compartments that are not easily accessible. To fill this gap, we investigated tissue-homing properties of cells that transiently appear in the circulating blood. Through cell separation and in vitro stimulation, the HIV-1 "Gag and Envelope reactivation co-detection assay" (GERDA) enables sensitive detection of Gag+/Env+ protein-expressing cells down to about one cell per million using flow cytometry. By associating GERDA with proviral DNA and polyA-RNA transcripts, we corroborate the presence and functionality of HIV-1 in critical body compartments utilizing t-distributed stochastic neighbor embedding (tSNE) and density-based spatial clustering of applications with noise (DBSCAN) clustering with low viral activity in circulating cells early after diagnosis. We demonstrate transcriptional HIV-1 reactivation at any time, potentially giving rise to intact, infectious particles. With single-cell level resolution, GERDA attributes virus production to lymph-node-homing cells with central memory T cells (T$_{CM}$s) as main players, critical for HIV-1 reservoir eradication.

Human immunodeficiency virus type 1 (HIV-1) infection is treated with antiretroviral therapy (ART), usually consisting of 2-3 different drugs, referred to as combination ART (cART). Our recent randomized clinical trial comparing a switch to dolutegravir monotherapy with continuation of cART in early-treated individuals demonstrated sustained virological suppression over 48 weeks. Here, we characterize the longitudinal landscape of the HIV-1 reservoir in these participants, with particular attention to potential differences between treatment groups regarding evidence of evolution as a proxy for low-level replication. Near full-length HIV-1 proviral polymerase chain reaction and next-generation sequencing was applied to longitudinal peripheral blood mononuclear cell samples to assess proviral evolution and the potential emergence of drug resistance mutations (DRMs). Neither an increase in genetic distance nor diversity over time was detected in participants of both treatment groups. Single proviral analysis showed high proportions of defective proviruses and low DRM numbers. No evidence for evolution during dolutegravir monotherapy was found in these early-treated individuals.

Background: Dolutegravir monotherapy (DTG-m) results in virological failure (VF) in some people with human immunodeficiency virus (PWH). We sought to identify the independent factors associated with the risk of VF and to explore the effect size heterogeneity between subgroups of PWH enrolled in DTG-m trials. Methods: We searched for randomized clinical trials (RCTs) evaluating DTG-m versus combined antiretroviral therapy (cART) among PWH virologically controlled for at least 6 months on cART. We performed an individual participant data meta-analysis of VF risk factors and quantified their explained heterogeneity in random-effect models. Definition of VF was a confirmed plasma human immunodeficiency virus (HIV)-1 ribonucleic acid (RNA) >50 copies/mL by week 48. Results: Among 416 PWH from 4 RCTs, DTG-m significantly increased the risk of VF (16 of 227 [7%] versus 0 of 189 for cART; risk difference 7%; 95% confidence interval [CI], 1%-2%; P = .02; I2 = 51%). Among 272 participants exposed to DTG-m, VF were more likely in participants with the following: first cART initiated ≥90 days from HIV acute infection (adjusted hazard ratio [aHR], 5.16; 95% 95% CI, 1.60-16.65), CD4 T cells nadir <350/mm3 (aHR, 12.10; 95% CI, 3.92-37.40), HIV RNA signal at baseline (aHR, 4.84; 95% CI, 3.68-6.38), and HIV-deoxyribonucleic acid (DNA) copy number at baseline ≥2.7 log/106 peripheral blood mononuclear cells (aHR, 3.81; 95% CI, 1.99-7.30). Among these independent risk factors, the largest effect size heterogeneity was found between HIV DNA subgroups (I2 = 80.2%; P for interaction = .02). Conclusions: Our study supports the importance of a large viral reservoir size for explaining DTG-m simplification strategy failure. Further studies are needed to link size and genetic diversity of the HIV-1 reservoir.

PURPOSE OF REVIEW HIV-1 drug resistance (HIV DR) testing is routinely performed by genotyping plasma viruses using Sanger population sequencing. Next-generation sequencing (NGS) is increasingly replacing standardized Sanger sequencing. This opens up new opportunities, but also brings challenges. RECENT FINDINGS The number of NGS applications and protocols for HIV DR testing is increasing. All of them are noninferior to Sanger sequencing when comparing NGS-derived consensus sequences to Sanger sequencing-derived sequences. In addition, NGS enables high-throughput sequencing of near full-length HIV-1 genomes and detection of low-abundance drug-resistant HIV-1 variants, although their clinical implications need further investigation. Several groups have defined remaining challenges in implementing NGS protocols for HIV-1 resistance testing. Some of them are already being addressed. One of the most important needs is quality management and consequently, if possible, standardization. SUMMARY The use of NGS technologies on HIV DR testing will allow unprecedented insights into genomic structures of virus populations that may be of immediate relevance to both clinical and research areas such as personalized antiretroviral treatment. Efforts continue to tackle the remaining challenges in NGS-based HIV DR testing.

Viruses are the cause of a considerable burden to human, animal and plant health, while on the other hand playing an important role in regulating entire ecosystems. The power of new sequencing technologies combined with new tools for processing "Big Data" offers unprecedented opportunities to answer fundamental questions in virology. Virologists have an urgent need for virus-specific bioinformatics tools. These developments have led to the formation of the European Virus Bioinformatics Center, a network of experts in virology and bioinformatics who are joining forces to enable extensive exchange and collaboration between these research areas. The EVBC strives to provide talented researchers with a supportive environment free of gender bias, but the gender gap in science, especially in math-intensive fields such as computer science, persists. To bring more talented women into research and keep them there, we need to highlight role models to spark their interest, and we need to ensure that female scientists are not kept at lower levels but are given the opportunity to lead the field. Here we showcase the work of the EVBC and highlight the achievements of some outstanding women experts in virology and viral bioinformatics.