A novel TaqMan real-time PCR assay to estimate ex vivo human immunodeficiency virus type 1 fitness in the era of multi-target (pol and env) antiretroviral therapy.
Despite numerous studies on human immunodeficiency virus type 1 (HIV-1) fitness, many key conceptual and technical questions are still unsolved. For example, the proper system to determine virus fitness of HIV-1 is still unknown. In this study, an assay was developed to estimate HIV-1 fitness based on growth competition experiments and TaqMan real-time PCR. This novel technique was compared with several methods (i.e. virus growth kinetics, growth competition/heteroduplex-tracking analysis and single-cycle replication capacity assay) in order to analyse the impact of various genomic regions and overall genetic background on virus fitness. HIV-1 primary isolates and three different sets of recombinant viruses [i.e. recombinant clones carrying protease (PR), reverse transcriptase (RT) or the 3' end of Gag, PR and RT (3'Gag/PR/RT), sequences amplified by PCR from the same primary isolates)] were evaluated. Here, it is demonstrated that, in spite of intrinsic differences, both growth competition/TaqMan and single-cycle replication assays detected a significant reduction in HIV-1 fitness as a consequence of drug-resistant mutations in pol. However, this new assay, based on HIV-1 isolates, may be useful to quantify replicative fitness in viruses from patients treated simultaneously with antiretroviral drugs targeting different genomic regions of HIV-1 (e.g. pol and env).
Weber
Jan
J
Department of Virology, Lerner Research Institute, Cleveland Clinic Foundation, NN10, 9500 Euclid Avenue, Cleveland, OH 44195, USA.
Rangel
Hector R
HR
Department of Virology, Lerner Research Institute, Cleveland Clinic Foundation, NN10, 9500 Euclid Avenue, Cleveland, OH 44195, USA.
Chakraborty
Bikram
B
Department of Virology, Lerner Research Institute, Cleveland Clinic Foundation, NN10, 9500 Euclid Avenue, Cleveland, OH 44195, USA.
Tadele
Mahlet
M
Department of Virology, Lerner Research Institute, Cleveland Clinic Foundation, NN10, 9500 Euclid Avenue, Cleveland, OH 44195, USA.
Martinez
Miguel A
MA
Laboratori de Retrovirologia, Fundacio irsiCaixa, Hospital Universitari Germans Trias I Pujol, Badalona, Spain.
Martinez-Picado
Javier
J
Laboratori de Retrovirologia, Fundacio irsiCaixa, Hospital Universitari Germans Trias I Pujol, Badalona, Spain.
Marotta
Michael L
ML
Department of Virology, Lerner Research Institute, Cleveland Clinic Foundation, NN10, 9500 Euclid Avenue, Cleveland, OH 44195, USA.
Mirza
Muneer
M
Department of Virology, Lerner Research Institute, Cleveland Clinic Foundation, NN10, 9500 Euclid Avenue, Cleveland, OH 44195, USA.
Ruiz
Lidia
L
Laboratori de Retrovirologia, Fundacio irsiCaixa, Hospital Universitari Germans Trias I Pujol, Badalona, Spain.
Clotet
Bonaventura
B
Laboratori de Retrovirologia, Fundacio irsiCaixa, Hospital Universitari Germans Trias I Pujol, Badalona, Spain.
Wrin
Terri
T
ViroLogic Inc., South San Francisco, CA, USA.
Petropoulos
Christos J
CJ
ViroLogic Inc., South San Francisco, CA, USA.
Quiñones-Mateu
Miguel E
ME
Department of Virology, Lerner Research Institute, Cleveland Clinic Foundation, NN10, 9500 Euclid Avenue, Cleveland, OH 44195, USA.
D019380:Anti-HIV Agents / Q000494:pharmacology*; D024882:Drug Resistance, Viral / Q000235:genetics; D004359:Drug Therapy, Combination; D015686:Gene Products, env / Q000235:genetics; D015684:Gene Products, pol / Q000235:genetics; D015658:HIV Infections / Q000150:complications / Q000188:drug therapy / Q000821:virology; D015497:HIV-1 / Q000145:classification / Q000187:drug effects / Q000254:growth & development* / Q000502:physiology*; D006801:Humans; D016133:Polymerase Chain Reaction / Q000379:methods*; D011995:Recombination, Genetic; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*; D019914:Taq Polymerase / Q000378:metabolism*; D014779:Virus Replication
D019380:Anti-HIV Agents; D015686:Gene Products, env; D015684:Gene Products, pol; D018894:Reverse Transcriptase Inhibitors; D019914:Taq Polymerase