Background
HIV-1 persistence poses a significant barrier to HIV eradication. Findings from the largest post-mortem study (FIND 2.0), which utilized minimally invasive tissue sampling in 38 virally suppressed individuals on antiretroviral therapy (ART) (VS; <400 copies/mL) and 22 unsuppressed (VU) untreated individuals showed the presence of proviral DNA in all 11 anatomically distinct compartments. We report detailed genotypic characterization of full length HIV-1 envelope glycoprotein (env) from this cohort.
Methods
Full length HIV-1 env was PCR amplified from extracted DNA from bone marrow aspirate (BM) and trephine (BMT), brain, bronchoalveolar lavage (BAL), cervicovaginal lavage, kidney, lymph nodes, liver, lung, spleen and whole blood (WB) from 60 FIND 2.0 decedents. Amplicons were sequenced (population-based) on Illumina MiSeq and analysed for subtype, minority variants, premature stop codons, hypermutations, intact rev response element (RRE), and coreceptor tropism. Viral compartmentalization was assessed by comparing sequence identity, coreceptor tropism prediction, V3 loop variants and phylogenetic clustering of each env per compartment/decedent.
Results
Of 521 DNA samples, 242 (46%) env amplified yielding 218 quality sequences: 127 from 36 VS, 91 from 19 VU. Phylogenetic clustering confirmed all were infected with HIV-1 subtype C, with sequences clustering by patient. Higher proportions of env sequences from VS were hypermutated (19%) as compared to VU (7%), with WB, BM, and BMT harboring most defective env. RRE was intact in 64% of VS and 52% of VU env sequences with most intact RREs detected in BAL. Premature stop codons were present in 24% of sequences, classifying them as defective. Minority variants were detected in 81% of env, with greater diversity in VU decedents. CCR5-tropic viruses predominated in both groups (92% VS; 80% VU), while CXCR4-tropic variants were more common in VU decedents, particularly in brain, liver and WB. Minor variant V3 loop sequence analysis showed mixtures of CCR5/CXCR4-utilizing viruses in some compartments, while single tropism sequences were found in others. Compartmentalization was observed in 40% VS and 63% VU decedents.
Conclusions
These findings offer new insights into the dynamic and complex proviral HIV-1 populations across distinct anatomical compartments, largely independent of ART status. We highlight the persistence of diverse, transcriptionally intact HIV-1 env in these anatomical sites with evidence of compartmentalization.
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