A15 Archived ART resistance in the latent reservoir of virally suppressed Ugandans

The increased access of antiretroviral therapy (ART) has drastically improved the health of infected individuals. However, increased levels of ART resistance globally threaten ART effectiveness. Resistance monitoring is currently limited to viremic individuals or prior to ART initiation. The archival nature of the HIV latent reservoir (LR) of virally suppressed patients allows examination for the persistence of ART-resistant latent viral variants. Whole blood samples were collected longitudinally in Rakai, Uganda, from 70 virally suppressed HIV-1 infected individuals. The quantitative viral outgrowth assay was performed to measure the frequency of replication-competent latently infected resting-memory CD4 + (rCD4) T-cells. RNA was extracted from HIV p24-positive outgrowth supernatant, and the reverse transcriptase (RT) region was sequenced using a validated site-specific next-generation sequencing assay (Illumina, San Diego, CA). Consensus sequences containing >2.5 per cent of the total raw amplicons of each outgrowth well were analyzed for ART drug resistance mutations using the Stanford Database. The presence of clonal sequence is expressed as both percent clonality and Shannon Entropy. Replication-competent virus was cultured from 52/70 (74.3%) individuals, of which, RT-pol sequence data were obtained from 49/52 (94.3%) individuals. The presence of ART-resistant virus was found in the LR from one individual on second-line therapy that included a protease inhibitor. There were 20 and 44 total prominent consensus sequences from all wells at years 1 and 3 of follow-up, respectively. ART-resistant mutations for both RT-inhibitor drug classes were found in 30 per cent and 27.3 per cent of the total prominent consensus sequences of this one individual from years 1 and 3, respectively. The major ART resistance profile in this individual included: M184V, Y188L, K191E, and G190A. The percentage of total outgrowth that was clonal (percent clonality) increased from year 1 to year 3 (38.1–81.8%) and Shannon Entropy decreased (0.722–0.576). The presence of archived replication-competent ART-resistant virus in the LR was found in only one individual. There were two ART-resistant prominent consensus sequences isolated at year 3 that were not sampled 2 years earlier. The persistence of resistant, intact replication-competent proviral sequences in the LR of this individual seem to be supported by clonal expansion.