588 Impact of Shorter Sequence Length on HIV-1 Drug Resistance Analysis Outcome and Clade Assignment via Virtual Phenotype P. Lecocq *1, M. Van Houtte 1, J. Veldeman 1, L. Bacheler 2 1VIRCO BVBA, Mechelen, Belgium and 2VIRCO USA, Durham, NC
Background:VirtualPhenotype provides a quantitative assessment of anti-HIV-1 drug susceptibility from a viral genotype. The basis of this assessment is an extensive database of genotypes and corresponding drug susceptibility phenotypes obtained from HIV-1 clinical isolates. The system as developed is based on all codons (1-99) of the protease gene and codons 1-400 of the reverse transcriptase (RT) gene, covering all positions with known resistance-associated mutations. The purpose of the current study was to assess the impact of use of shorter sequence lengths on accuracy of phenotypic resistance analysis and on clade determination.
Methods: Mutations at positions 318 and 333 of RT are known to affect susceptibility to NNRTIs and NRTIs, respectively. Full-length sequences extracted from the database were truncated to retain codons 1-99 of protease and 40-247 or 1-320 of RT, thus generating sequence lengths corresponding to those generated by other HIV-1 genotyping systems. VirtualPhenotype analyses of distinct genotypes involving positions 318 (n = 98) and 333 (n = 343) of RT were compared for truncated vs full-length sequences. For assessment of clade attribution, both a new alignment tool based on ClustalW algorithm to generate alignment vs LANL 79 HIV-1 subtype references and a new clade determination system (iterative sliding windows) were used.
Results: Changes in susceptibility prediction (resistant to susceptible or vice-versa) resulted when some genotypes with mutations 318 or 333 were truncated. Given the prevalence of those genotypes in the current database of clinical isolates, susceptibility prediction changed for 0.02% (efavirenz and nevirapine)-0.12% (delavirdine) of isolates with mutation Y318F, and for 0.33% (ZDV) of isolates with mutations G333E or D. A dataset of 1,000 sequences was used to test the efficiency of the enhanced clade assessment. The effect of truncated sequence length on clade assessment was negligible (< 0.1%).
Conclusions: Given the current prevalence of clinical isolates with known resistance-associated mutations in codons 1-400 of RT, the impact of truncated sequence lengths on resistance analysis was limited to fewer than 0.5% of clinical isolates. Changes in prevalence of known mutations in this region, or identification/selection of new resistance-associated mutations could affect this conclusion in the future. Clade assessment was minimally affected by the use of the shortened sequences.