Frequency matters: comparison of drug resistance mutation detection by Sanger and next-generation sequencing in HIV-1

• Published in: Journal of antimicrobial chemotherapy 2023-03, Vol.78 (3), p.656-664
• Main Authors: Balakrishna, Suraj, Loosli, Tom, Zaheri, Maryam, Frischknecht, Paul, Huber, Michael, Kusejko, Katharina, Yerly, Sabine, Leuzinger, Karoline, Perreau, Matthieu, Ramette, Alban, Wymant, Chris, Fraser, Christophe, Kellam, Paul, Gall, Astrid, Hirsch, Hans H, Stoeckle, Marcel, Rauch, Andri, Cavassini, Matthias, Bernasconi, Enos, Notter, Julia, Calmy, Alexandra, Günthard, Huldrych F, Metzner, Karin J, Kouyos, Roger D
• Format: Article
• Language: English
• Subjects: Anti-HIV Agents - therapeutic use; Cohort Studies; Drug Resistance, Viral - genetics; Genotype; High-Throughput Nucleotide Sequencing - methods; HIV Infections - drug therapy; HIV Seropositivity - drug therapy; HIV-1; Humans; Male; Mutation; Viral Load
• ISSN: 0305-7453; 1460-2091
• DOI: 10.1093/jac/dkac430

Abstract Background Next-generation sequencing (NGS) is gradually replacing Sanger sequencing (SS) as the primary method for HIV genotypic resistance testing. However, there are limited systematic data on comparability of these methods in a clinical setting for the presence of low-abundance drug resistance mutations (DRMs) and their dependency on the variant-calling thresholds. Methods To compare the HIV-DRMs detected by SS and NGS, we included participants enrolled in the Swiss HIV Cohort Study (SHCS) with SS and NGS sequences available with sample collection dates ≤7 days apart. We tested for the presence of HIV-DRMs and compared the agreement between SS and NGS at different variant-calling thresholds. Results We included 594 pairs of SS and NGS from 527 SHCS participants. Males accounted for 80.5% of the participants, 76.3% were ART naive at sample collection and 78.1% of the sequences were subtype B. Overall, we observed a good agreement (Cohen’s kappa >0.80) for HIV-DRMs for variant-calling thresholds ≥5%. We observed an increase in low-abundance HIV-DRMs detected at lower thresholds [28/417 (6.7%) at 10%–25% to 293/812 (36.1%) at 1%–2% threshold]. However, such low-abundance HIV-DRMs were overrepresented in ART-naive participants and were in most cases not detected in previously sampled sequences suggesting high sequencing error for thresholds