HIV reservoir quantification using cross-subtype multiplex ddPCR

Noah A J Cassidy; Carolyn S Fish; Claire N 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

doi:10.1016/j.isci.2021.103615

HIV reservoir quantification using cross-subtype multiplex ddPCR

iScience. 2021 Dec 11;25(1):103615. doi: 10.1016/j.isci.2021.103615. eCollection 2022 Jan 21.

Authors

Noah A J Cassidy¹, Carolyn S Fish¹, Claire N Levy², Pavitra Roychoudhury^{3

4}, Daniel B Reeves³, Sean M Hughes², Joshua T Schiffer^{3

5}, Sarah Benki-Nugent⁶, Grace John-Stewart^{6

5

7

8}, Dalton Wamalwa⁹, Keith R Jerome^{3

4}, Julie Overbaugh¹, Florian Hladik^{2

3

5}, Dara A Lehman^{1

6}

Affiliations

¹ Division of Human Biology, Fred Hutchinson Cancer Research Center, Seattle, WA, USA.
² Department of Obstetrics and Gynecology, University of Washington, Seattle, WA, USA.
³ Vaccine and Infectious Disease Division, Fred Hutchinson Cancer Research Center, Seattle, WA, USA.
⁴ Department of Laboratory Medicine and Pathology, University of Washington, Seattle, WA, USA.
⁵ Department of Medicine, University of Washington, Seattle, WA, USA.
⁶ Department of Global Health, University of Washington, Seattle, WA, USA.
⁷ Department of Epidemiology, University of Washington, Seattle, WA, USA.
⁸ Department of Pediatrics, University of Washington, Seattle, WA, USA.
⁹ Department of Pediatrics and Child Health, University of Nairobi, Kenyatta National Hospital, Nairobi, Kenya.

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.

Keywords: Genotyping; Virology.

Abstract

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