Andrew J. Leigh Brown

HIV-1 transmission precipitates a stringent genetic bottleneck, with 75% of new infections initiated by a single genetic variant. Where multiple variants initiate infection, recipient set point viral load (SpVL) and the rate of CD4 ⁺ T cell decline may be elevated, but these findings remain inconsistent. Here, we summarised the evidence for this phenomenon, then tested whether previous studies possessed sufficient statistical power to reliably identify a true effect of multiple variant infection leading to higher SpVL. Next, we combined models of HIV-1 transmission, heritability and disease progression to understand whether available data suggest a faster CD4 ⁺ T cell decline would be expected to associated with multiple variant infection, without an explicit dependency between the two. First, we found that most studies had insufficient power to identify a true significant difference, prompting an explanation for previous inconsistencies. Next, our model framework revealed we would not expect to observe a positive association between multiple variant infections and faster CD4 ⁺ T cell decline, in the absence of an explicit dependency. Consequently, while empirical evidence may be consistent with a positive association between multiple variant infection and faster CD4 ⁺ T cell decline, further investigation is required to establish a causal basis.

HIV incidence in Kazakhstan increased by 73% between 2010 and 2020, with an estimated 35,000 people living with HIV (PLHIV) in 2020. The development of antiretroviral drug resistance is a major threat to effective antiretroviral therapy (ART), yet studies on the prevalence of drug resistance in Kazakhstan are sparse. In this study on the molecular epidemiology of HIV in Kazakhstan, we analyzed 968 partial HIV-1 pol sequences that were collected between 2017 and 2020 from PLHIV across all regions of Kazakhstan, covering almost 3% of PLHIV in 2020. Sequences predominantly represented subtypes A6 (57%) and CRF02_AG (41%), with 32% of sequences exhibiting high-level drug resistance. We further identified distinct drug-resistant mutations (DRMs) in the two subtypes: subtype A6 showed a propensity for DRMs A62V, G190S, K101E, and D67N, while CRF02_AG showed a propensity for K103N and V179E. Codon usage analysis revealed that different mutational pathways for the two subtypes may explain the difference in G190S and V179E frequencies. Phylogenetic analysis highlighted differences in the timing and geographic spread of both subtypes within the country, with A62V-harboring subtype A6 sequences clustering on the phylogeny, indicative of sustained transmission of the mutation. Our findings suggest an HIV epidemic characterized by high levels of drug resistance and differential DRM frequencies between subtypes. This emphasizes the importance of drug resistance monitoring within Kazakhstan, together with DRM and subtype screening at diagnosis, to tailor drug regimens and provide effective, virally suppressive ART.

The Sustainable East Africa Research in Community Health (SEARCH) trial was a universal test-and-treat (UTT) trial in rural Uganda and Kenya, aiming to lower regional HIV-1 incidence. Here, we quantify breakthrough HIV-1 transmissions occurring during the trial from population-based, dried blood spot samples. Between 2013 and 2017, we obtained 549 gag and 488 pol HIV-1 consensus sequences from 745 participants: 469 participants infected prior to trial commencement and 276 SEARCH-incident infections. Putative transmission clusters, with a 1.5% pairwise genetic distance threshold, were inferred from maximum likelihood phylogenies; clusters arising after the start of SEARCH were identified with Bayesian time-calibrated phylogenies. Our phylodynamic approach identified nine clusters arising after the SEARCH start date: eight pairs and one triplet, representing mostly opposite-gender linked (6/9), within-community transmissions (7/9). Two clusters contained individuals with non-nucleoside reverse transcriptase inhibitor (NNRTI) resistance, both linked to intervention communities. The identification of SEARCH-incident, within-community transmissions reveals the role of unsuppressed individuals in sustaining the epidemic in both arms of a UTT trial setting. The presence of transmitted NNRTI resistance, implying treatment failure to the efavirenz-based antiretroviral therapy (ART) used during SEARCH, highlights the need to improve delivery and adherence to up-to-date ART recommendations, to halt HIV-1 transmission.

Objective: To describe the use of a postarrest debriefing tool (DBT) within a university teaching hospital and to evaluate user perceptions of the tool.Design: Observational study over a 1‐year period and associated hospital clinical personnel survey.Setting: University teaching hospital.Interventions: Qualitative data surrounding the use and utility of the DBT were analyzed, as well as survey results.Measurements and Main Results: Forty‐four arrests occurred during the study period. Debriefing was performed after 26 of 44 (59%) cardiopulmonary resuscitation (CPR) events, of which 22 of 26 (85%) were recorded using the DBT and four without the DBT. Return of spontaneous circulation did not significantly affect the use of the DBT (p = 0.753). Most events in which debriefing was not performed occurred outside of business hours (13/18; 72%). The most frequent positive debriefing comments related to cooperation/coordination within the team (22/167; 13%). The most frequent negative debriefing comments concerned equipment issues (36/167; 22%). Of the action points generated, 57% (34/60) were directed at equipment use/availability. Teams reported that emergency drugs were appropriately administered in 21 of 22 (95%) cases. In contrast, closed loop communication was reportedly only used during 6 of 22 (27%) events.The hospital survey response rate was 56 of 338 (17%) clinical staff, of whom 37 of 56 (66%) agreed or strongly agreed that debriefing had improved team performance during CPR. Overall, 33 of 56 (60%) staff felt that the DBT had improved the debriefing process at the hospital. However, 3 of 56 (5%) staff members felt that they were unable to state their opinions in a blame‐free environment during debriefing.Conclusions: Implementation of a DBT enabled formal identification of strengths and training needs of resuscitation teams, and its implementation was viewed positively by the majority of hospital staff. However, further refinement of the tool and prospective studies evaluating its efficacy in improving outcome are warranted.

Summary: Background: HIV-1 infections initiated by multiple founder variants are characterised by a higher viral load and a worse clinical prognosis, yet little is known about the routes of exposure through which transmission of multiple founder variants is most likely. Methods: We conducted a systematic review of studies that estimated founder variant multiplicity in HIV-1 infection, searching MEDLINE, EMBASE and Global Health databases for papers published between 1 ^st January 1990 and 14 ^th September 2020 (PROSPERO study CRD42020202672 ). Leveraging individual patient estimates from these studies, we performed a logistic meta-regression to estimate the probability that an HIV infection is initiated by multiple founder variants. We calculated a pooled estimate using a random effects model, subsequently stratifying this estimate across nine transmission routes in a univariable analysis. We then extended our model to adjust for different study methods in a multivariable analysis, recalculating estimates across the nine transmission routes. Findings: We included 70 publications in our analysis, comprising 1657 individual patients. Our pooled estimate of the probability that an infection is initiated by multiple founder variants was 0·25 (95% CI: 0·21-0·29), with moderate heterogeneity ( Q = 132 · 3, p < 0 · 001, I ² = 64 · 2%). Our multivariable analysis uncovered differences in the probability of multiple variant infection by transmission route. Relative to a baseline of male-to-female transmission, the predicted probability for female-to-male multiple variant transmission was significantly lower at 0·13 (95% CI: 0·08-0·20), while the probabilities for people-who-inject-drugs (PWID) and men-who-have-sex-with-men (MSM) transmissions were significantly higher at 0·37 (0·24-0·53) and 0·30 (0·33-0·40), respectively. There was no significant difference in the probability of multiple variant transmission between male-to-female transmission (0·21 (0·14-0·31)), post-partum mother-to-child (0·18 (0·03-0·57)), pre-partum mother-to-child (0·17 (0·08-0·33)), intrapartum mother-to-child (0·27 (0·14-0·40)). Interpretation: We identified PWID and MSM transmissions are significantly more likely to result in an infection initiated by multiple founder variants, whilst female-to-male infections are significantly less likely. Quantifying how the routes of HIV infection impact the transmission of multiple variants allows us to better understand how the evolution and epidemiology of HIV-1 determine clinical outcomes. Funding: This study was supported by the MRC Precision Medicine Doctoral Training Programme (ref: 2259239) and an ERC Starting Grant awarded to KEA (award number 757688). The funding sources played no role in study design, data collection, data analysis, data interpretation, or writing of the report. Panel: Research in context: Evidence before this study: Most HIV-1 infections are initiated by a single, genetically homogeneous founder variant. Infections initiated by multiple founders, however, are associated with a significantly faster decline of CD4+ T cells in untreated individuals, ultimately leading to an earlier onset of AIDS. Through our systematic search of MEDLINE, EMBASE and Global Health databases, we identified 82 studies that classify the founder variant multiplicity of early HIV infections. As these studies vary in the methodology used to calculate the number of founder variants, it is difficult to evaluate the multiplicity of founder variants across routes of exposure. Added value of this study: We estimated the probability that an HIV infection is initiated by multiple founder variants across exposure routes, leveraging individual patient data from 70 of the identified studies. Our multivariable meta-regression adjusted for heterogeneity across study methodology and uncovered differences in the probability that an infection is initiated by multiple founder variants by exposure route. While overall, we estimated that 25% of infections are initiated by multiple founder variants, our analysis found that this probability for female-to-male transmission is significantly lower than for male-to-female transmission. By contrast, this probability was significantly higher among people-who-inject-drugs (PWID) and men-who-have-sex-with-men (MSM). There was no difference in the probability of multiple founder variant transmission for mother-to-child transmission when compared with male-to-female sexual transmission. Implications of all the available evidence: Because HIV-1 infections initiated by multiple founders are associated with a poorer prognosis, determining whether the route of exposure affects the probability with which infections are initiated by multiple variants facilitates an improved understanding of how the evolution and epidemiology of HIV-1 determine clinical progression. Our results identify that PWID and MSM transmissions are significantly more likely to result in an infection initiated by multiple founder variants compared to male-to-female. This reiterates the need for focussed public health programmes that reduce the burden of HIV-1 in these risk groups.

Phylogenetic inference is useful in characterising HIV transmission networks and assessing where prevention is likely to have the greatest impact. However, estimating parameters that influence the network structure is still scarce, but important in evaluating determinants of HIV spread. We analyzed 2017 HIV pol sequences (728 Lake Victoria fisherfolk communities (FFCs), 592 female sex workers (FSWs) and 697 general population (GP)) to identify transmission networks on Maximum Likelihood (ML) phylogenetic trees and refined them using time-resolved phylogenies. Network generative models were fitted to the observed degree distributions and network parameters, and corrected Akaike Information Criteria and Bayesian Information Criteria values were estimated. 347 (17.2%) HIV sequences were linked on ML trees (maximum genetic distance ≤4.5%, ≥95% bootstrap support) and, of these, 303 (86.7%) that consisted of pure A1 (n = 168) and D (n = 135) subtypes were analyzed in BEAST v1.8.4. The majority of networks (at least 40%) were found at a time depth of ≤5 years. The waring and yule models fitted best networks of FFCs and FSWs respectively while the negative binomial model fitted best networks in the GP. The network structure in the HIV-hyperendemic FFCs is likely to be scale-free and shaped by preferential attachment, in contrast to the GP. The findings support the targeting of interventions for FFCs in a timely manner for effective epidemic control. Interventions ought to be tailored according to the dynamics of the HIV epidemic in the target population and understanding the network structure is critical in ensuring the success of HIV prevention programs.

The General Population Cohort (GPC) in south-western Uganda has a low HIV-1 incidence rate (25 years (aOR = 1.52; 95% CI, 1.16–2.0) and being a resident in the GPC (aOR = 6.90; 95% CI, 5.22–9.21). Phylogeographic analysis showed significant viral dissemination (Bayes Factor test, BF > 3) from the GPC without significant viral introductions (BF < 3) into the GPC. The findings suggest localized HIV-1 transmission in the GPC. Intensifying geographically focused combination interventions in the GPC would contribute towards controlling HIV-1 infections.

Recombination is an important feature of HIV evolution, occurring both within and between the major branches of diversity (subtypes). The Ugandan epidemic is primarily composed of two subtypes, A1 and D, that have been co-circulating for 50 years, frequently recombining in dually infected patients. Here, we investigate the frequency of recombinants in this population and the location of breakpoints along the genome. As part of the PANGEA-HIV consortium, 1,472 consensus genome sequences over 5 kb have been obtained from 1,857 samples collected by the MRC/UVRI & LSHTM Research unit in Uganda, 465 (31.6 per cent) of which were near full-length sequences (>8 kb). Using the subtyping tool SCUEAL, we find that of the near full-length dataset, 233 (50.1 per cent) genomes contained only one subtype, 30.8 per cent A1 (n = 143), 17.6 per cent D (n = 82), and 1.7 per cent C (n = 8), while 49.9 per cent (n = 232) contained more than one subtype (including A1/D (n = 164), A1/C (n = 13), C/D (n = 9); A1/C/D (n = 13), and 33 complex types). K-means clustering of the recombinant A1/D genomes revealed a section of envelope (C2gp120-TMgp41) is often inherited intact, whilst a generalized linear model was used to demonstrate significantly fewer breakpoints in the gag–pol and envelope C2-TM regions compared with accessory gene regions. Despite similar recombination patterns in many recombinants, no clearly supported circulating recombinant form (CRF) was found, there was limited evidence of the transmission of breakpoints, and the vast majority (153/164; 93 per cent) of the A1/D recombinants appear to be unique recombinant forms. Thus, recombination is pervasive with clear biases in breakpoint location, but CRFs are not a significant feature, characteristic of a complex, and diverse epidemic.

Background: Memory CD4+ T cells (mCD4s) containing integrated HIV DNA are considered the main barrier to a cure for HIV infection. Here we analysed HIV DNA reservoirs in antigen-specific subsets of mCD4s to delineate the mechanisms by which HIV reservoirs persist during ART. Methods/Results: HIV Gag, Cytomegalovirus (CMV) and Tetanus Toxoid (TT) specific mCD4s were isolated from peripheral blood samples obtained from 11 individual subjects, 2-11 years after commencing ART. Antigen-specific mCD4s were identified by the sensitive OX40 assay and purified by cell sorting. Total HIV DNA levels were quantified by real-time PCR, and clonal viral sequences generated from mCD4 subsets and pre-ART plasma samples. Quantitative results and sequence analysis were restricted to 5 and 3 study participants respectively, likely due to the low frequency of the antigen-specific mCD4s and relatively low HIV DNA proviral loads. Median HIV Gag-, CMV-, and TT-specific mCD4s were 0.61%, 2.46% and 0.78% of total mCD4s, and contained a median of 2.50, 2.38, and 2.55 log10 copies of HIV DNA per 106 cells respectively. HIV DNA sequences derived from antigen-specific mCD4s clustered with sequences derived from pre-ART plasma samples. There was a trend towards increased viral diversity in clonal viral sequences derived from CMV-specific relative to TT-specific mCD4s. Discussion/Conclusions: Despite limitations, this study provides direct evidence that HIV reservoirs persist in memory CD4+ T cell subsets maintained by homeostatic proliferation (TT) and adds to growing evidence against viral evolution during ART. Similar future studies require techniques that sample diverse HIV reservoirs and with improved sensitivity.