Simple detection of point mutations associated with HIV-1 drug resistance

A novel assay is described For the detection of HIV-1 drug resistance that is simple, cheap and sensitive. HIV-1 drug resistance in B and non-B HIV-1 subtypes was investigated using Mutagenically-Separated PCR (MSPCR) - a com petitive semi-nested PCR which uses mutagenic primers. The assay was assess ed for sensitivity. specificity and its ability to detect mutant virus with in a mixed mutant-wild-type population, Gene sequencing was carried out sim ultaneously for comparison. MS-PCR detected five copies of HIV-1 RNA from l aboratory isolates and 50 copies from patient samples. We demonstrate 100% specificity of detection for wild type or mutant virus for clades A, B, C, D and E. For mixed populations of virus, MS-PCR can detect at least a 10% m ix of wild type:mutant, or vice-versa. When applied to African patient samp les MS-PCR detected 91.6% of the codons tested. Concordance with sequencing data was 88.8% for protease and 97.2% for RT. MS-PCR is sensitive and spec ific for the detection of mutations in HIV-1. and can be adapted easily to test for resistance at any codon of interest. (C) 2001 Elsevier Science B.V . All rights reserved.