A novel quantitative approach for measuring the reservoir of latent HIV-1 proviruses

A stable latent reservoir for HIV-1 in resting CD4(+) T-cells precludes cure(1–3). Curative strategies targeting the reservoir are being tested(4,5) and require accurate, scalable reservoir assays. The reservoir was defined with quantitative viral outgrowth assays (QVOAs) for cells releasing infectious virus following one round of T-cell activation(1). However, QVOAs and newer assays for cells producing viral RNA after activation(6) may underestimate reservoir size because one round of activation does not induce all proviruses(7). Many studies rely on simple PCR-based assays to detect proviral DNA regardless of transcriptional status, but the clinical relevance of these assays is unclear, as the vast majority proviruses are defective(7–9). We describe a novel approach that separately quantifies intact and defective proviruses and show that the dynamics of cells carrying intact and defective proviruses are different in vitro and in vivo, a finding with implications for targeting the intact proviruses that are a barrier to cure.