HIV-1 Genetic Variation in Infected Individuals

Division Of Basic Sciences - Nci

Investigators

Linked publications & trials

Abstract

HIV replication in vivo is rapid and error prone, and results in large and genetically diverse populations of HIV-infected cells. We are using population-based sequencing, as well as the single-genome sequencing (SGS) technology we developed previously. In addition, with M. Kearney's group (HIV DRP), we have investigated the use of next-generation sequencing (NGS) approaches (Illumina sequencing) to study HIV population genetics. We have utilized these approaches to analyze and understand the accumulation of genetic variation in gag/pol and env. We have made significant advances in additional assay development and have extended studies to a number of different patient groups, including chronically infected patients, both naive and on therapy, as well as in primary and early HIV infection. As a result, we are obtaining a more comprehensive picture of HIV genetic variation in vivo in the presence or absence of drug resistance. _____We have previously reported that replicating population size is substantial prior to initiating antiretroviral therapy (ART). We have applied NGS approaches to refine the estimates of HIV and found that 1X107 infected cells may be replicating in infected individuals daily. In our new studies, we have been able to investigate rates of recombination in vivo and found that recombination occurs at a high rate in vivo, such that there are very few linked polymorphisms in any virus population, and that recombination represents a potentially important mechanism for spread of HIV mutations. As resistance to integrase inhibitors increases, and NIH clinics are enrolling more such patients, we are studying the development of resistance to this important class of HIV inhibitors. ____We have recently reported the emergence of high-level resistance to dolutegravir with the addition of a single secondary mutation at position T97A. We have now completed a detailed analysis of the emergence of resistance and found that the T97A variant emerged from a limited number (3-4) of variants that arose rapidly by replication and, as predicted by our population genetics studies, through recombination. The development of these techniques has led to new insights in HIV population dynamics in understanding the effects of ART, the nature of replication in natural suppression of HIV, and population dynamics of non-subtype B HIV populations. ____The dynamics of HIV-1 populations in patients undergoing ART remain uncertain, and we are conducting an extensive genetic analysis of HIV-1 before and after initiation of ART (completed Protocol 97-I-0082, new Protocol 08-I-0221). These results will yield new information regarding the nature and timing of genetic bottlenecks occurring during ART. Analysis of HIV-1 sequences at relatively low viremia has been limited by technical issues in amplifying the relatively few HIV-1 sequences present in plasma during therapy. We have successfully adapted the SGS procedure to obtain acceptable numbers of sequences from patients suppressed on ART. In collaboration with M. Polis and D. Persaud (NIH Bench to Bedside Award, 2006), we are analyzing genetic variation in patients enrolled in Protocol 97-I-0082 (now 08-I-0221; F. Maldarelli, PI) who have been suppressed on ART for prolonged (greater than 8 y) periods. Initial analyses demonstrate that HIV does not undergo a genetic bottleneck upon initiation of ART; despite a 100- to 10,000-fold decline in levels of peripheral viremia, no significant decreases in genetic diversity were detected in the first 1-2 y of therapy. These data indicate a common source of virus infecting short-lived cells (responsible for greater than 90-99% of virus produced prior to therapy) and longer-lived cells (responsible for virus produced 1-2 years after therapy is initiated). After prolonged therapy, emergence of predominant clones (as previously noted by Bailey et al.) was detected in the majority (7/8) patients. We are also applying population genetics approaches to quantify the emergence of drug-resistance mutations in rebound viremia in patients undergoing ART. We are specifically investigating the relative roles of mutation and selection in development of resistance to AZT and NNRTI, as well as quantifying the role of APOBEC-mediated mutations in the emergence of the M184I mutation conferring resistance to 3TC, FTC, and, to a degree, abacavir. ____We are using these approaches to investigate the spread of HIV in the US and in selected populations in the world. We have demonstrated clear evidence of early spread of drug-resistant HIV in the HIV epidemic in Washington, DC, which has the highest prevalence of HIV in the US. We are now investigating the spread of HIV in DC using samples stored early in the epidemic. We have demonstrated that subtype C HIV, the most common subtype in the world, was present a number of years earlier than previously thought, and prior to the rapid expansion of this subtype in sub-Saharan Africa. These studies have yielded information and new sequences that are critical to understanding the current spread of HIV in DC and will inform public health strategies to eliminate the spread of HIV in DC. In addition, we recently reported analysis of a mini-epidemic spread of a specific subtype A HIV variant among intravenous drug users (IVDU) associated with cathinone abuse in Israel. Despite public health efforts to halt the spread, we found this variant was continuing to spread in IVDU and could also be detected in other HIV risk groups. Understanding the expansion of genetic diversity following infection from a genetically limited to a highly diverse population has useful implications for applicability in understanding the HIV epidemic. Based on our understanding of genetic variation in acute and chronically infected individuals, we developed a new bioinformatics algorithm to discriminate between recently and chronically infected individuals based exclusively on population-based commercial genotyping data. Development of this algorithm has yielded the invention report EIR #238-2009. This approach has been used in a variety of settings, and we have used it to investigate spread of HIV in DC and more recently in analysis of the subtype A mini-epidemic in Tel Aviv, Israel. ____These studies have shed new light on the spread of HIV in populations and inform the public health efforts to eradicate HIV worldwide.