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.