HIV reservoir quantification using cross-subtype multiplex ddPCR
Noah A.J. Cassidy, Carolyn S. Fish, Claire Levy, Pavitra Roychoudhury, Daniel B. Reeves, Sean M. Hughes, Joshua T. Schiffer, Sarah Benki‐Nugent, Grace John‐Stewart, Dalton Wamalwa, Keith R. Jerome, Julie Overbaugh, Florian Hladik, Dara A. Lehman
Abstract
A major barrier to conducting HIV cure research in populations with the highest HIV burden is the lack of an accurate assay to quantify the replication-competent reservoir across the dominant global HIV-1 subtypes. Here, we modify a subtype B HIV-1 assay that quantifies both intact and defective proviral DNA, adapting it to accommodate cross-subtype HIV-1 sequence diversity. We show that the cross-subtype assay works on subtypes A, B, C, D, and CRF01_AE and can detect a single copy of intact provirus. In longitudinal blood samples from Kenyan infants infected with subtypes A and D, patterns of intact and total HIV DNA follow the decay of plasma viral load over time during antiretroviral therapy, with intact HIV DNA comprising 7% (range 1%-33%) of the total HIV DNA during HIV RNA suppression. This high-throughput cross-subtype reservoir assay will be useful in HIV cure research in Africa and Asia, where HIV prevalence is highest.