Convergent Evolution of Reverse Transcriptase (RT) Genes of Human Immunodeficiency Virus Type 1 Subtypes E and B following Nucleoside Analogue RT Inhibitor Therapies

Abstract

Changes in the drug susceptibility, gene lineage, and deduced amino acid sequences of the reverse transcriptase (RT) of human immunodeficiency virus type 1 (HIV-1) subtype E following 3-azido-3-deoxythymidine (AZT) monotherapy or AZT-2,3-dideoxyinosine combination therapy were examined with sequential virus isolates from a single family. The changes were compared to those reported for HIV-1 subtype B, revealing striking similarities in selected phenotype and amino acids independent of differences in the RT backbone sequences that constantly distinguish the two subtypes. Particularly, identical amino acid substitutions were present simultaneously at four different positions (D67N, K70R, T215F, and K219Q) for high-level AZT resistance. These data suggest that HIV-1 subtypes E and B evolve convergently at the phenotypic and amino acid levels when the nucleoside analogue RT inhibitors act as selective forces.

Augmentation of Human Immunodeficiency Virus Type 1 Subtype E (CRF01_AE) Multiple-Drug Resistance by Insertion of a Foreign 11-Amino-Acid Fragment into the Reverse Transcriptase

“…As expected from the previous clinical, genetic, and phenotypic data (35,36), 93JP-NH3 was as sensitive to the inhibitors tested as reference HIV-1 strain NL43 was (Fig. 1B, 93JP-NH3; the fold increases in IC 50 ranged from 1.0 to 1.8).…”

“…Molecular cloning of a full-length HIV-1 subtype E proviral genome was carried out as described previously for HIV-1 subtype B (37). Briefly, a single restriction enzyme site (SpeI) in the HIV-1 subtype E 93JP-NH1 (35,36) genome was identified by Southern blot analysis of the unintegrated circular DNAs prepared by the Hirt method (13) from 93JP-NH1-infected MT2 cells. Subsequently, the circular DNAs were digested with SpeI, purified on the basis of their sizes (9 to 12 kb), ligated with SpeI-digested arms of the ZAP Express lambda phage vector (Stratagene), and packaged in vitro with Gigapack III Gold packaging extract (Stratagene).…”

“…Peripheral blood was taken from the patient in December 1999 (99JP-NH3-II), and the pol gene segment (1,035 bp) of the plasma viral RNA (99JP-NH3-IIp) and the RNA of the viruses isolated in culture (99JP-NH3-IIvm) were sequenced by direct sequencing. The deduced amino acid sequences were aligned with a subtype E consensus derived from early 1990s samples in Thailand (23), along with the sequence obtained in 1993 from NH3 (36).…”

Isolation and Molecular Characterization of a Nelfinavir (NFV)-Resistant Human Immunodeficiency Virus Type 1 That Exhibits NFV-Dependent Enhancement of Replication

“…The method used to infect cells has been described previously (23)(24)(25). Briefly, PHA-stimulated PBMCs (2 ϫ 10 5 cells), MT-2 cells (2 ϫ 10 4 cells), and PM-1 cells (2 ϫ 10 4 cells) were infected with 0.2 ml of cell-free supernatant containing HIV-1 (2 ϫ 10 5 32 P cpm of RT activity) in the absence or presence of NFV (0.1 and 1 M) at 37°C for 16 h, washed once, and cultured in 0.2 ml of culture medium with the same concentration of NFV.…”

Phylogenetically resolving epidemiologic linkage

“…Thus, we evaluated trees as if they were fully resolved, perfectly sampled, and generated by the neutral coalescence processes described by our within-host model. In reality, short branches may not be resolved due to lack of mutations, all relevant lineages may not be sampled, and phylogenetic uncertainty can also occur with large amounts of homoplasy induced by convergent selection or recombination (47)(48)(49)(50)(51). In general, fast-evolving genomic regions, such as env, have been shown to accumulate enough mutations to robustly recover branches (28).…”