Relative replication fitness of a high-level 3 '-azido-3 '-deoxythymidine-resistant variant of human immunodeficiency virus type 1 possessing an amino acid deletion at codon 67 and a novel substitution (Thr -> Gly) at codon 69

The combination of an amino acid deletion at codon 67 (Delta 67) and Thr-to -Gly change at codon 69 (T69G) in the reverse transcriptase (RT) of human i mmunodeficiency virus type 1 (HIV-1) is associated with high-level resistan ce to multiple RT inhibitors. To determine the relative contributions of th e Delta 67 and T69G mutations on viral fitness, we performed a series of st udies of HIV replication using recombinant variants. A high-level 3 ' -azid o-3 ' -deoxythymidine (AZT)-resistant variant containing Delta 67 plus T69G /K70R/L741/KI03N/T215F/ K219Q in RT replicated as efficiently as wild-type virus (Wt). In contrast, the construct without Delta 67 exhibited impaired replication (23% of growth of Wt). A competitive fitness study failed to re veal any differences in replication rates between the Delta 67+T69G/K70R/L7 41/KI03N/T215F/K219Q mutant and Wt. Evaluation of proviral DNA sequences ov er a 3-year period in a patient harboring the multiresistant HIV revealed t hat the T69G mutation emerged in the context of a D67NiK70R/T215F/K219Q mut ant backbone prior to appearance of the Delta 67 deletion. To assess the im pact of this stepwise accumulation of mutations on viral replication, a ser ies of recombinant variants was constructed and analyzed for replication co mpetence. The T69G mutation was found to confer 2 ' ,3 ' -dideoxyinosine re sistance at the expense of fitness. Subsequently, the development of the De lta 67 deletion led to a virus with improved replication and high-level AZT resistance.