Naive vs memory CD4 T cells: which subset harbors the real reservoir?

Background: Despite the efficacy of combination antiretroviral therapy (cART) to reduce plasma HIV RNA to undetectable levels, the virus persists in a latent form and remains a hurdle to eradication. CD4+ T lymphocytes harbor the majority of the HIV reservoir, but the role of each CD4 T subset remains unclear. Methods: To explore the composition and character of the reservoir in CD4 T cell subsets, we sorted CD4 T cells into central, transitional, effector memory and naïve T cells (Tn). We measured HIV DNA levels and performed proviral sequencing of more than 1,400 proviruses in two subjects at an early and late time point after cART initiation. The contribution of each subset to the reservoir was calculated. We also examined proviral phylogeny to study the role of cellular differentiation in HIV persistence. Results: Tn contributed a surprisingly high percentage of intact proviruses to the reservoir (35% and 58% for subject 1 and 2), as compared to memory cells. The HIV integration level in Tn was > 10-fold lower than in memory T cells. The sequences isolated from Tn were mostly unique while the proviral populations in the effector memory were mainly clonal. Moreover, an UpSet analysis showed that the number of clones increased as cells differentiated from naïve to central memory to effector memory. Conclusions: Tn appear to be a major contributor to the intact reservoir in two HIV infected individuals. This contribution was maintained longitudinally two and nine years after ART. Our data suggest Tn and Tcm repopulate the memory reservoir over time.