The relevance of ultradeep sequencing for low HIV‐1 viral loads and proviruses in the clinical setting

Abstract

Improving the therapeutic management of HIV‐positive persons is a major public health issue and includes better detection of drug resistance mutations (DRMs). The aim of this study was (i) to explore DRMs in HIV‐1‐positive persons presenting a blood viral load (VL) < 1000 genomes copies (gc)/mL, including the analyze of cerebrospinal fluid (CSF) and HIV‐DNA from peripheral blood mononuclear cells using ultradeep sequencing (UDS) and (ii), to evaluate how these DRMs could influence the clinical practices. For each patient (n = 12), including five low‐VL patients (i.e., <1000 gc/mL), HIV‐1 UDS targeting the protease, reverse transcriptase and integrase genes was performed on plasma, proviral DNA, and CSF when available. Sequencing discordances or failures were mostly found in samples from low‐VL patients. A 5% UDS cut‐off allowed to increase the sensitivity to detect DRMs in different compartments, excepted in CSF. Patients with the highest viral quasispecies heterogeneity were naïve of treatment or presented a medical history suggesting low selection pressure or virological failures. When analyzing compartmentalization and following‐up patients: low‐frequency variants (LFVs) were responsible for 47% (n = 8) and 76% (n = 13) of changes in drug resistance interpretation, respectively. In such cases, we conclude that UDS is a robust technique, which still could be improved by increase the RNA and/or DNA extraction in low‐VL samples to detect LFVs. Further studies are needed to define the impact of LFVs on antiretroviral treatments. At last, when considering a DRM, the use of mutational load would probably be more suitable than frequencies.