Paired quantitative and qualitative assessment of the replication-competent HIV-1 reservoir and comparison with integrated proviral DNA

Julio C.C. Lorenzi, Yehuda Z. Cohen, Lillian B. Cohn, Edward F. Kreider, John P. Barton, Gerald H. Learn, Thiago Oliveira, Christy L. Lavine, Joshua A. Horwitz, Allison Settler, Mila Jankovic, Michael S. Seaman, Arup K. Chakraborty, Beatrice H. Hahn, Marina Caskey, Michel C. Nussenzweig

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HIV-1-infected individuals harbor a latent reservoir of infected CD4+ T cells that is not eradicated by antiretroviral therapy (ART). This reservoir presents the greatest barrier to an HIV-1 cure and has remained difficult to characterize, in part, because the vast majority of integrated sequences are defective and incapable of reactivation. To characterize the replication-competent reservoir, we have combined two techniques, quantitative viral outgrowth and qualitative sequence analysis of clonal outgrowth viruses. Leukapheresis samples from four fully ART-suppressed, chronically infected individuals were assayed at two time points separated by a 4- to 6-mo interval. Overall, 54% of the viruses emerging from the latent reservoir showed gp160 env sequences that were identical to at least one other virus. Moreover, 43% of the env sequences from viruses emerging from the reservoir were part of identical groups at the two time points. Groups of identical expanded sequences made up 54% of proviral DNA, and, as might be expected, the sequences of replication-competent viruses in the active reservoir showed limited overlap with integrated proviral DNA, most of which is known to represent defective viruses. Finally, there was an inverse correlation between proviral DNA clone size and the probability of reactivation, suggesting that replication-competent viruses are less likely to be found among highly expanded provirus-containing cell clones.

Original languageEnglish
Pages (from-to)E7908-E7916
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number49
StatePublished - 6 Dec 2016
Externally publishedYes

Bibliographical note

Funding Information:
We thank all study participants who devoted time to our research. We also thank members of the M.C.N. laboratory for helpful discussions. J.C.C.L. is supported by Award 248676/2013-0 from the Brazil Conselho Nacional de Pesquisas "Ciencia sem Fronteiras." Y.Z.C. is supported by Grant KL2TR001865, National Center for Advancing Translational Sciences (NCATS), and Grant UL1 TR000043, National Institutes of Health (NIH) Clinical and Translational Science Award (CTSA) program. J.C.C.L. and Y.Z.C. are additionally supported by NIH/National Institute of Allergy and Infectious Diseases Grant U01 AI118536 (to M.C.). The contributions of J.P.B. and A.K.C. were supported by the Ragon Institute of MGH, MIT, and Harvard. E.F.K., G.H.L., and B.H.H. were supported by BEAT-HIV Delaney Collaboratory Grant UM1 AI126620. M.C.N. is a Howard Hughes Medical Investigator.


  • Culture
  • HIV
  • Method
  • Replication-competent
  • Reservoir


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