Characterization of Genetic Variation and 3'-Azido-3'-Deoxythymidine- Resistance Mutations of Human Immunodeficiency Virus by the RNase A Mismatch Cleavage Method

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 88; no. 10; pp. 4280 - 4284
• Main Authors: Lopez-Galindez, Cecilio, Rojas, Jose M., Najera, Rafael, Richman, Douglas D., Perucho, Manuel
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences of the United States of America 15.05.1991; National Acad Sciences
• Subjects: Acquired Immunodeficiency Syndrome - microbiology; AIDS/HIV; Base Sequence; Codon; Codons; DNA, Viral - genetics; Drug Resistance, Microbial - genetics; Genes, Viral; Genetic mutation; Genetic Variation; HIV; HIV - drug effects; HIV - genetics; HIV Envelope Protein gp120 - genetics; Humans; Molecular Sequence Data; Mutation; Nucleic Acid Hybridization; Nucleotides; Phenotypes; Polymerase Chain Reaction; Ribonuclease, Pancreatic; RNA; RNA probes; Viral Proteins - genetics; Viruses; Zidovudine - pharmacology
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.88.10.4280

The RNase A mismatch cleavage method has been applied to the characterization of natural genetic variation of human immunodeficiency virus (HIV) from different geographical areas. The approach provides a rapid and simple assay for the analysis of differences in closely related viral isolates and allows the establishment of phylogenetic relationships between epidemiologically distinct viruses. Our results show a broad clustering of circulating viruses according to their geographical distribution. We also have analyzed the temporal appearance of mutations associated with the acquisition of resistance to 3'-azido-3'-deoxythymidine (AZT). The results show that mutations in codon 215 of the viral reverse transcriptase can be detected readily by this method in HIV isolates and also directly in peripheral blood from HIV-infected individuals after in vitro amplification of viral sequences with the polymerase chain reaction. The specific recurrence of identical double-nucleotide substitutions in epidemiologically and geographically distant viruses suggests that the restricted amino acid substitutions at this position selected by drug exposure are a critical, rate-limiting step in the acquisition of drug resistance.