Biology subjects

Kirkby, D.

Publications and source records attributed to Kirkby, D..

3 recordsLinked to original sources

bioRxiv2026

HIV superinfection reveals sequential reservoir reactivation and immune-driven rebound dynamics

Superinfection, where a person with HIV acquires a second phylogenetically distinct strain, provides an opportunity to study reservoir evolutionary dynamics because the two strains can be tracked over time. We here characterize such a case that originally went undetected by clinical monitoring. The participant, who expressed the protective HLA-B*57:03 allele, initially controlled his subtype B infection but lost control after superinfection with a unique recombinant form (URF), which came to dominate in plasma without displacing the original B strain. Five years after initiating therapy, replication-competent proviruses from both strains persisted in blood, though URF proviruses dominated (80%). Despite their unequal reservoir representation, both strains rapidly rebounded upon treatment interruption, though most rebounding sequences belonged to a URF subclade that predated ART initiation. During the treatment interruption, genetically distinct viral lineages continually appeared in plasma, consistent with sequential "waves" of reactivation of diverse reservoir clones. Notably, immune escape variants emerged from the reservoir in a later "wave", displacing initial susceptible variants and underscoring the important role of immune responses in shaping rebound dynamics. Sequence analysis revealed that the URF transmitted/founder virus already harbored key HLA-B*57:03-associated escape mutations, likely explaining the loss of viral control after superinfection. Our results reveal that superinfection can remain undetected despite routine HIV clinical monitoring, that pre-adaptation of the superinfecting strain to host HLA can undermine immune control, that rebound virus can originate from reservoir clones predating ART initiation, and that immune responses can actively shape rebound dynamics by driving escape variant outgrowth during treatment interruption. IMPORTANCEThis case demonstrates that HIV superinfection can go undetected by routine clinical monitoring, underscoring the importance of assays that capture HIV diversity -- particularly those used to screen participants for cure trials. It also illustrates how pre-adaptation of a superinfecting strain to host HLA-restricted immune responses can evade existing immunity, with implications for understanding natural immune control, and underscoring the challenge of viral diversity for vaccine and immunotherapy design. The sequential emergence of genetically diverse HIV lineages during treatment interruption, including minority immune escape variants that emerged from the reservoir to displace the initial susceptible population, highlights the difficulty of predicting rebound virus composition and underscores the key role of immune responses in shaping rebound dynamics. Critically, escaped rebound variants can then reseed the reservoir, thereby enriching it in escaped forms, highlighting a specific vulnerability for cure approaches that harness natural immune responses to eliminate reservoir cells.

molecular biology↗

bioRxiv2023

HIV reservoirs are dominated by genetically younger and clonally enriched proviruses

In order to cure HIV, we need to better understand the within-host evolutionary origins of the small reservoir of genome-intact proviruses that persists within infected cells during antiretroviral therapy (ART). Most prior studies on reservoir evolutionary dynamics however did not discriminate genome-intact proviruses from the vast background of defective ones. We reconstructed within-host pre-ART HIV evolutionary histories in six individuals and leveraged this information to infer the ages of intact and defective proviruses sampled after an average >9 years on ART, along with the ages of rebound and low-level/isolated viremia occurring during this time. We observed that the longest-lived proviruses persisting on ART were exclusively defective, usually due to large deletions. In contrast, intact proviruses and rebound HIV exclusively dated to the years immediately preceding ART. These observations are consistent with genome-intact proviruses having shorter lifespans, likely due to the cumulative risk of elimination following viral reactivation and protein production. Consistent with this, intact proviruses (and those with packaging signal defects) were three times more likely to be genetically identical compared to other proviral types, highlighting clonal expansion as particularly important in ensuring their survival. By contrast, low-level/isolated viremia sequences were genetically heterogeneous and sometimes ancestral, where viremia may have originated from defective proviruses. Results reveal that the HIV reservoir is dominated by clonally-enriched and genetically younger sequences that date to the untreated infection period when viral populations had been under within-host selection pressures for the longest duration. Knowledge of these qualities may help focus strategies for reservoir elimination. ImportanceCharacterizing the HIV reservoir that endures despite antiretroviral therapy (ART) is critical to cure efforts. Our observation that the oldest proviruses persisting during ART were exclusively defective, while intact proviruses (and rebound HIV) all dated to the years immediately pre- ART, explains why prior studies that sampled sub-genomic proviruses on-ART (which are largely defective) routinely found sequences dating to early infection, whereas those that sampled viral outgrowth sequences found essentially none. Together with our findings that intact proviruses were also more likely to be clonal, and that on-ART low-level/isolated viremia originated from proviruses of varying ages (including possibly defective ones), our observations indicate that: 1) on-ART and rebound viremia can have distinct within-host origins, 2) intact proviruses have shorter lifespans than grossly-defective ones, and therefore depend on clonal expansion for persistence, and 3) the HIV reservoir, being overall genetically younger, will be substantially adapted to within-host pressures, complicating immune-based cure strategies.

genetics↗

bioRxiv2020

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

Opening Paragraph (serves as abstract for submission) and BodyThe Intact Proviral DNA Assay (IPDA) was developed to address the critical need for a precise and scalable method for intact HIV reservoir quantification1. This duplexed droplet digital PCR (ddPCR) assay simultaneously targets the HIV Packaging Signal ({Psi}) and the Rev Responsive Element (RRE) within Envelope (env) to distinguish genomically intact proviruses against a large background of defective ones2. 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 studies4-7. 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.

genetics↗