Development of an assay that detects transcriptionally competent human immunodeficiency virus type one particles.
To study the functional properties of HIV-1 reverse transcriptase (RT) from intact viral particles without the requirement for tissue culture expansion, a method that couples HIV-1 reverse transcription utilizing its endogenous RT (ERT) with polymerase chain reaction amplification (PCR) was developed. Detection of endogenous reverse transcripts from HIV particles by ERT-PCR was compared to HIV RNA PCR detection using avian myeloblastosis virus (AMV) RT from plasma samples from 45 HIV-1 infected patients. The HIV ERT-PCR method was capable of detecting plasma viremia with the same efficiency (29/29 patients) as the AMV RT HIV RNA PCR in patients with CD4 cell counts of less than 500/mm3. The determination of HIV-RT drug sensitivities using four well-characterized HIV-1 lab strains was assessed. The ERT-PCR method detected reduced sensitivity to TIBO R82150 (10 microM) in a TIBO resistant strain but not in the TIBO sensitive HTLV-IIIB viral mixture or an HTLV-IIIB clone. In summary, the HIV ERT-PCR method provides a useful approach for the detection of HIV and the characterization of RT sensitivities among HIV-1 strains.
Busso
M
M
Retrovirology Research Department, Mount Sinai Medical Center, Miami Beach, FL 33140.
Resnick
L
L
D000998:Antiviral Agents / Q000494:pharmacology; D001569:Benzodiazepines / Q000494:pharmacology; D004279:DNA, Viral / Q000235:genetics; D054306:Dideoxynucleotides; D004352:Drug Resistance, Microbial; D054303:HIV Reverse Transcriptase; D015497:HIV-1 / Q000201:enzymology / Q000302:isolation & purification*; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D016133:Polymerase Chain Reaction / Q000379:methods*; D012194:RNA-Directed DNA Polymerase / Q000097:blood* / Q000235:genetics; D018894:Reverse Transcriptase Inhibitors; D012680:Sensitivity and Specificity; D013942:Thymine Nucleotides / Q000494:pharmacology; D015215:Zidovudine / Q000031:analogs & derivatives / Q000494:pharmacology
D000998:Antiviral Agents; D004279:DNA, Viral; D054306:Dideoxynucleotides; D007093:Imidazoles; D018894:Reverse Transcriptase Inhibitors; D013942:Thymine Nucleotides; C062687:R 82150; D001569:Benzodiazepines; D015215:Zidovudine; C052873:zidovudine triphosphate; D054303:HIV Reverse Transcriptase; D012194:RNA-Directed DNA Polymerase