HIV-1 APOBEC-context Mutations Detected by Next-Generation Sequencing in Plasma: Implications for Drug Resistance Interpretation

Abstract

BACKGROUND & AIMS: APOBEC3-mediated cytidine deamination produces characteristic G?A substitutions in HIV genomes, classically described in proviral DNA but also detectable in plasma HIV-1 RNA using next-generation sequencing (NGS). APOBEC-signature mutations reflect enrichment of G?A substitutions in APOBEC target motifs, whereas APOBEC-context mutations correspond to substitutions at drug resistance-associated positions compatible with APOBEC editing. We evaluated their prevalence and implications for resistance interpretation in plasma NGS sequences. METHODS: We performed a retrospective multicentre analysis of plasma HIV-1 NGS-sequences from antiretroviral-naïve individuals in the period 2022-2023. Sequencing was performed in participating laboratories, and FASTQ files were centrally reanalysed using DeepChek. APOBEC-signature mutations, APOBEC-context drug resistance mutations (DRMs), and stop codons were evaluated across protease, reverse transcriptase, and integrase. Variant frequencies were analysed using thresholds of 1%, 3%, and 5%. RESULTS: Among 290 individuals, 268 had complete NGS data. At a 1% threshold, APOBEC-signature mutations were detected in 159 individuals and APOBEC-context DRMs in 64 (22%). The coexistence of both patterns was observed in 52 individuals (18%), decreasing to 17 (6%) and 8 (3%) at 3% and 5% thresholds, respectively. Certain mutations, including M184I, G190E, G140S, and R263K, remained detectable at higher thresholds. Integrase mutation R263K frequently co-occurred with multiple APOBEC-signature mutations and stop codons, consistent with extensive APOBEC-mediated editing. CONCLUSIONS: APOBEC-associated mutational patterns can be detected in plasma HIV-1 RNA by NGS and may affect interpretation of resistance results. Conservative frequency thresholds (5%) and evaluation of APOBEC signatures may help reduce misclassification of APOBEC-induced variants as clinically relevant resistance.