Human immunodeficiency virus type 1 protease inhibitor drug-resistant mutants give discordant results when compared in single-cycle and multiple-cycle fitness assays.
The replication fitness of HIV-1 drug-resistant mutants has been measured using either multiple-cycle or single-cycle assays (MCAs or SCAs); these assays have not been systematically compared. We developed an MCA and an SCA that utilized either intact or env-deleted recombinant viral vectors, respectively, in which virus-infected cells were detected by flow cytometry of a reporter gene product. Fitness was measured using each assay for 11 protease mutants, 9 reverse transcriptase mutants, and two mutants with mutations in gag p6, which is important for the release of virus particles from the cell membrane. In the SCA, fitness (replication capacity [RC]) was defined as the proportion of cells infected by the mutant compared to the wild type 40 h after infection. MCA fitness (1+s) was determined by comparing the changes in the relative proportions of cells infected by the mutant and the wild type between 3 and 5 days after infection. Five protease mutants showed statistically different fitness values by the MCA versus the SCA: the D30N, G48V, I50V, I54L, and I54M mutants. When all the mutants were ranked in order from most to least fit for both assays, 4 protease mutants moved more than 5 positions in rank: the D30N, I54L, I54M, and V82A mutants. There were no significant differences in fitness for the gag p6 or reverse transcriptase mutants. We propose that discordant results in the MCA and SCA are due to alterations in late events in the virus life cycle that are not captured in an SCA, such as burst size, cell-to-cell transmission, or infected-cell life span.
Dykes
Carrie
C
Department of Medicine, University of Rochester School of Medicine and Dentistry, Rochester, New York 14642, USA. carrie_dykes@urmc.rochester.edu
Wu
Hulin
H
Sims
Matthew
M
Holden-Wiltse
Jeanne
J
Demeter
Lisa M
LM
D024882:Drug Resistance, Viral; D015658:HIV Infections / Q000821:virology*; D016333:HIV Protease / Q000235:genetics*; D017320:HIV Protease Inhibitors / Q000494:pharmacology*; D015497:HIV-1 / Q000187:drug effects* / Q000502:physiology*; D006801:Humans; D014773:Virology / Q000379:methods; D014774:Virulence; D014779:Virus Replication
D017320:HIV Protease Inhibitors; D016333:HIV Protease; C514803:p16 protease, Human immunodeficiency virus 1
Proc Natl Acad Sci U S A. 1991 Apr 15;88(8):3195-9
2014240
J Virol. 2005 Jul;79(14):8979-90
15994792
Virology. 2004 Sep 1;326(2):262-79
15302212
J Virol. 2010 Aug;84(15):7581-91
20504921
J Virol. 1995 Jun;69(6):3712-20
7745720
Virology. 2006 Sep 15;353(1):184-92
16797050
J Virol. 2006 Mar;80(5):2380-9
16474144
J Virol. 2002 Nov;76(21):11104-12
12368352
J Virol. 1998 May;72(5):3773-8
9557659
J Virol. 2001 Oct;75(20):9644-53
11559796
J Virol. 2000 Oct;74(20):9339-46
11000201
J Virol. 2007 Mar;81(6):3037-41
17192300
J Acquir Immune Defic Syndr. 2007 Aug 1;45(4):411-7
17554216
J Virol. 1996 Apr;70(4):2581-5
8642689
Proc Natl Acad Sci U S A. 1994 Sep 27;91(20):9564-8
7937806
J Virol. 2004 Feb;78(4):1718-29
14747537
Clin Microbiol Rev. 2007 Oct;20(4):550-78
17934074
J Virol. 2009 Jan;83(1):140-9
18971283
J Clin Microbiol. 2006 Jun;44(6):1930-43
16757582
Trends Microbiol. 2003 Feb;11(2):56-9
12598123
Eur J Biochem. 1999 Jul;263(1):238-45
10429209
J Virol. 2003 Sep;77(18):10172-5
12941931
J Virol. 2000 Sep;74(18):8390-401
10954539
Antimicrob Agents Chemother. 2000 Apr;44(4):920-8
10722492
J Virol. 2000 Sep;74(18):8524-31
10954553
J Virol. 1999 Jul;73(7):5803-13
10364332
Biochemistry. 2003 Dec 30;42(51):15029-35
14690411
J Biol Chem. 2004 Jan 2;279(1):509-16
14551187
J Virol. 1997 Aug;71(8):5871-7
9223476
AIDS. 2002 May 3;16(7):1009-17
11953467
Gene. 1985;40(1):1-8
3912262
J Virol. 1999 May;73(5):3744-52
10196268
Science. 1995 Jan 27;267(5197):483-9
7824947
J Virol. 2004 May;78(9):4628-37
15078945
J Virol. 2007 May;81(9):4540-50
17287264
J Virol. 2003 Nov;77(22):12105-12
14581547
Virology. 2006 May 10;348(2):462-74
16504235
Proc Natl Acad Sci U S A. 1996 Dec 24;93(26):15266-71
8986799
Antivir Chem Chemother. 2002 Jan;13(1):27-37
12180647
Proc Natl Acad Sci U S A. 1996 Oct 15;93(21):11395-9
8876146
Proc Natl Acad Sci U S A. 1996 Feb 20;93(4):1648-53
8643685
Rev Med Virol. 1997 Jul;7(2):87-96
10398474
J Virol. 2008 Jan;82(1):495-502
17942528
Science. 1989 Mar 31;243(4899):1731-4
2467383
J Virol. 1991 Jan;65(1):162-9
1845882
J Virol. 2004 Jan;78(2):603-11
14694092
J Infect Dis. 2004 Jul 15;190(2):251-6
15216458
Virology. 2001 Jul 5;285(2):193-203
11437654
J Acquir Immune Defic Syndr. 2001 May 1;27(1):7-13
11404514
Science. 1985 Aug 9;229(4713):563-6
2992081
Biochemistry. 2002 Jul 9;41(27):8613-9
12093278