HIV Diversity Considerations in the Application of the Intact Proviral DNA Assay (IPDA)

Opening Paragraph (serves as abstract for submission) and Body The Intact Proviral DNA Assay (IPDA) was developed to address the critical need for a precise and scalable method for intact HIV reservoir quantification 1 . This duplexed droplet digital PCR (ddPCR) assay simultaneously targets the HIV Packaging Signal (Ψ) and the Rev Responsive Element (RRE) within Envelope ( env ) to distinguish genomically intact proviruses against a large background of defective ones 2 . The IPDA requires less time, resources, and biological material than the gold standard for replication-competent HIV reservoir measurement, the Quantitative Viral Outgrowth Assay (QVOA) 3 , and is being adopted in research and clinical studies 4–7 . In our cohort of HIV-1 subtype B-infected individuals from North America however, the IPDA yielded a 28% failure rate due to HIV polymorphism. We further demonstrate that within-host HIV diversity can lead the IPDA to underestimate intact HIV reservoir size, which could negatively impact clinical trial results interpretation. While the IPDA represents an important methodological advance, HIV diversity should be addressed before its widespread adoption.