Multimethod Longitudinal HIV Drug Resistance Analysis in Antiretroviral-Therapy-Naive Patients

Abstract

The global intensification of antiretroviral therapy (ART) can lead to increased rates of HIV drug resistance (HIVDR) mutations in treated and also in ART-naive patients. ART-naive HIV-1-infected patients from Cameroon were subjected to a multimethod HIVDR analysis using amplification-refractory mutation system (ARMS)-PCR, Sanger sequencing, and longitudinal next-generation sequencing (NGS) to determine their profiles for the mutations K103N, Y181C, K65R, M184V, and T215F/Y. We processed 66 ART-naive HIV-1-positive patients with highly diverse subtypes that underlined the predominance of CRF02_AG and the increasing rate of F2 and other recombinant forms in Cameroon. We compared three resistance testing methods for 5 major mutation sites. Using Sanger sequencing, the overall prevalence of HIVDR mutations was 7.6% (5/66) and included all studied mutations except K65R. Comparing ARMS-PCR with Sanger sequencing as a reference, we obtained a sensitivity of 100% (5/5) and a specificity of 95% (58/61), caused by three false-positive calls with ARMS-PCR. For 32/66 samples, we obtained NGS data and we observed two additional mismatches made up of minority variants (7% and 18%) that might not be clinically relevant. Longitudinal NGS analyses revealed changes in HIVDR mutations in all five positive subjects that could not be attributed to treatment. In one of these cases, superinfection led to the temporary masking of a resistant virus. HIVDR mutations can be sensitively detected by ARMS-PCR and sequencing methods with comparable performances. Longitudinal changes in HIVDR mutations have to be considered even in the absence of treatment.

Near full genome characterization of HIV‐1 unique recombinant forms in Cameroon reveals dominant CRF02_AG and F2 recombination patterns

“…They were shipped from the MDC under regulatory guidelines of transfer on biological samples to NYUSoM, where plasma was separated from peripheral blood mononuclear cells through Ficoll gradient centrifugation (Histopaque, Sigma‐Aldrich, St. Louis, MO, USA), and stored at −80°C. Eighteen URFs were selected based on previously published work from our group using partial genome sequencing (total of 509 participants) . Inclusion criteria were genetic evidence of URF infection and available plasma volumes ≥500 μL.…”

“…Possibly as a consequence, it has one of the most genetically diverse HIV epidemics in the world . The predominant lineage is the recombinant CRF02_AG, which accounts for more than half of infections . Also, circulating virus lineages include virtually every known HIV‐1 group M (HIV‐1 M) pure subtype, many CRFs, and a variety of URFs composed of pure, CRF and/or non‐classifiable sequences making this country a unique source of rare and emerging HIV‐1 M viral variants .…”

“…The predominant lineage is the recombinant CRF02_AG, which accounts for more than half of infections . Also, circulating virus lineages include virtually every known HIV‐1 group M (HIV‐1 M) pure subtype, many CRFs, and a variety of URFs composed of pure, CRF and/or non‐classifiable sequences making this country a unique source of rare and emerging HIV‐1 M viral variants . The range of subtypes and the extent of viral diversity within a geographical region considerably impact HIV diagnosis, treatment and prevention .…”

“…As part of our ongoing molecular surveillance efforts in Cameroon, our laboratories have in recent years identified and characterized two CRFs (CRF_36cpx and CRF_37cpx) , and we have monitored a steady increase in URFs . Through the application of a robust and versatile near full genome (NFG) amplification and sequencing technique , the current study characterized 18 near‐full genomes of HIV‐1 M recombinants.…”

Anti-V2 Antibody Deficiency in Individuals Infected With HIV-1

“…Glycoprotein 120 from 12 plasma viruses (6 each from the V2-deficient and V2-reactive groups) was sequenced, and virus subtypes were identified (Table 1). All but one virus (clade C) were HIV-1 circulating recombinant forms (CRF), which dominate in Cameroon, according to recent analysis with the most prevalent CRF02_AG (64.9%) and CRF22_01A1 (7.1%) (11, 12). The identified subtypes in this study were as follows: CRF02_AG (3 plasma samples), CRF01_AE (2), CRF22_01A1 (2), CRF11_cpx, CRF18_cpx, CRF36_cpx, and CRF37_cpx (Table 1).…”