Development and introduction of a new extended assay for resistance testing of HIV-1 into the Antiviral Resistance Testing Service.

PM Cook, SA Graham, E Smit HPA Birmingham, Dept. Pathology, Heartlands Hospital, Birmingham B9 5SS

HIV-1 drug susceptibility testing by means of genotyping has been carried out by the Antiviral Resistance Testing Unit in Birmingham since its inception in the 1990s. The unit developed an in-house assay which sequences the entire protease gene (Codons 1-99) and the first 235 codons of the reverse transcriptase (RT) gene in a nested PCR, to give a total of 1,000 nucleotide base pairs (bp) of sequence. Recently, a number of primary and accessory resistance mutations have been identified in areas that lie outside RT codon 235, including codons P236L, Y318L, N348I and G333D/E. A new extended assay for HIV-1 genotype resistance testing was constructed, covering the entire protease gene as before and the RT gene up to codon 360, generating a nested PCR product of 1857bp. Evaluation of the new extended assay showed it to be at least as sensitive as the original assay in that it amplified the same variety of diverse HIV-1 subtypes. Based on this the extended assay replaced the old assay by the end of 2008. Results of a comparison between the old assay and the first two months after the introduction of the extended assay into clinical practice will be presented.