Performance characteristcis of Allele-Specific PCR (ASPCR) in detecting drug resistance mutations among non-B HIV-1 Variants

J Virol Methods. 2023 Nov 22:114856. doi: 10.1016/j.jviromet.2023.114856. Online ahead of print.ABSTRACTAllele-Specific Polymerase Chain Reaction (ASPCR) is an affordable point-mutation assay whose validation could improve the detection of HIV-1 drug resistance mutations (DRMs) in resource-limited settings (RLS). We assessed the performance of ASPCR onforty-four non-B HIV-1 plasma samples from patients who were ARV treated in failure in N'Djamena-Chad. Viral RNA was reverse-transcribed and amplified using LightCycler® FastStart DNA MasterPLUS SYBR Green I. Detection of six major DRMs (K70R, K103N, Y181C, M184V, T215F, T215Y) was evaluated on Roche LightCycler®480 automated system (with dilutions 0.01-100%). ASPCR-results were compared to Sanger-sequencing (gold-standard). Correlations of mutation curves were excellent (R2>0.97); all DRMs were detected with desirable mutant/wild-type threshold differences (ΔCt≥9) except K70R(ΔCtK70R=6; ΔCtK103N=13; ΔCtM184V=9; ΔCtT215F=12; ΔCtT215Y=12; ΔCtY181C=9) and positive controls were below required thresholds. Also, ASPCR reproducibility on DRMs was assessed by using dilutions of intra-assay and inter-assay coefficient of variations respectively with a threshold of less than 50(i.e.<0.50 variation) which are;: K70R (0.02-0.28 vs. 0.12-0.37), K103N (0.08-0.42 vs. 0.12-0.37), Y181C (0.12-0.39 vs. 0.31-0.37), M184V (0.13-0.39 vs. 0.23-0.42), T215F (0.05-0.43 vs. 0.04-0.45) and T215Y (0.13-0.41 vs. 0.19-0.41). DRM detection...

Source: Journal of Virological Methods - November 24, 2023 Category: Virology Authors: Chatt é Adawaye Joseph Fokam Erick Ntambwe Kamangu Derrick Tambe Ayuk Ngwese Fabrice Susin Ali Mahamat Moussa BertinTchombou Hig-Zounet Joseph Mad-Toingu é Abdelsalam Tidjani Dolores Vaira Michel Moutschen Source Type: research