Intact HIV proviruses persist in children 7-9 years after initiation of ART in the first year of life

In adults starting ART during acute infection, 2% of proviruses that persist on ART are genetically intact by sequence analysis. By contrast, a recent report in early treated children failed to detect sequence intact proviruses. We sought to detect and characterize proviral sequences in another cohort of early treated children after 6-9 years on suppressive ART. PBMCs from perinatally-infected children from the CHER study were analysed. Near full-length proviral amplification and sequencing (NFL-PAS) was performed at one time point after 6-9 years on ART. Amplicons with large internal deletions were excluded (< 9kb). All amplicons =9kb were sequenced and analysed through a bio-informatic pipeline to detect indels, frameshifts or hypermutations that would render them defective. In eight children who started ART at a median age of 5.4 months (range: 2.0 - 11.1), 733 single NFL-PAS amplicons were generated. Of these, 534(72.9%) had large internal deletions, 174(23.7%) had hypermutation, 15(1.4%) had small internal deletions, 3(1.0%) had deletions in the packaging signal/major splice donor site and 7(1.0%) were sequence intact. These 7 intact sequences were from three children who initiated ART after 2.3 months of age; of whom one had two identical intact sequences, suggestive of a cell clone harbouring a replication-competent provirus. No intact proviruses were detected in four children who initiated ART before 2.3 months of age. Rare, intact proviruses can be detected in children who initiate ART after 2.3 months of age and are probably just as in adults, maintained by clonal expansion of cells infected before ART initiation.Importance There is limited data on the proviral landscape in early treated, long-term suppressed children, particularly in Sub-Saharan Africa where HIV-1 subtype C predominates. Investigating the sequence-intact reservoir could provide insight on the mechanisms by which intact proviruses persist and inform ongoing cure efforts. Through near full-length proviral amplification and sequencing (NFL-PAS) we generated 733 NFL-PAS amplicons from 8 children. We showed that rare, genetically intact proviruses could be detected in children who initiated ART after 2.3 months of age. The frequency of intact proviruses was lower than (p< 0.05) reported for HIV subtype B infected adults treated during early HIV infection. We show that cells harbouring genetically intact HIV proviruses are rare in early treated long-term suppressed children and may require the processing of a large number of cells to assess reservoir size. This points to the need for efficient methods to accurately quantify latent reservoirs particularly in paediatric studies where sample availability is limited. Copyright © 2019 American Society for Microbiology.

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