UpToDate
Official reprint from UpToDate®
www.uptodate.com ©2016 UpToDate®

Overview of HIV drug resistance testing assays

Author
Michael J Kozal, MD
Section Editor
Martin S Hirsch, MD
Deputy Editor
Jennifer Mitty, MD, MPH

INTRODUCTION

The use of drug resistance testing has become an integral part of HIV clinical care. The first clinical description of HIV resistance to antiretroviral drugs was published in 1989 about patients taking zidovudine monotherapy; accumulation of mutations within the reverse transcriptase gene resulted in a marked increase in drug resistance [1]. Subsequently, HIV variants resistant to every available antiretroviral agent have been identified in viral culture in the presence of drug and in treated HIV-infected patients. The evolution of drug resistance has significant clinical implications for choosing effective antiretroviral regimens [2].

Resistance assays can be categorized as either phenotypic or genotypic. These assays detect resistance in fundamentally different ways, although the results generally correlate with each other. There may be clinical settings in which one or the other assay could offer theoretical advantages or in which each assay would provide complementary information. However, the optimal specific use of these assays has not been definitively established.

The clinically available antiretroviral drug resistance assays will be discussed here, including the benefits and limitations of each type. Clinical trials evaluating these assays and the clinical use of these drug resistance assays in various patient populations are discussed elsewhere. (See "Clinical trials of HIV drug resistance testing" and "Drug resistance testing in the clinical management of HIV infection" and "Primer on interpretation of HIV drug resistance testing".)

PHENOTYPIC RESISTANCE ASSAYS

Phenotypic resistance assays measure the extent to which an antiretroviral drug inhibits virus replication in vitro. Similar to bacteriologic methods, this is typically performed by demonstrating an increase in the inhibitory concentration (IC) that is required to inhibit in vitro growth by 50 percent (IC50) compared with virus replication in the absence of drug. Results are reported as a fold-change in drug susceptibility of the patient sample compared with a laboratory reference strain.

Recombinant virus assay — Historically, phenotypic resistance assays were performed using virus cultured from patient peripheral blood mononuclear cells [3]. These assays, which required culturing virus in primary human peripheral blood mononuclear cells, were plagued by significant variability and lengthy turnaround times. They required approximately two to four weeks for initial HIV isolation, one week to titer the virus inoculum, and an additional one to two weeks to assay virus replication in the presence of different drug concentrations.

                           

Subscribers log in here

To continue reading this article, you must log in with your personal, hospital, or group practice subscription. For more information or to purchase a personal subscription, click below on the option that best describes you:
Literature review current through: Nov 2016. | This topic last updated: Tue Oct 07 00:00:00 GMT 2014.
The content on the UpToDate website is not intended nor recommended as a substitute for medical advice, diagnosis, or treatment. Always seek the advice of your own physician or other qualified health care professional regarding any medical questions or conditions. The use of this website is governed by the UpToDate Terms of Use ©2016 UpToDate, Inc.
References
Top
  1. Larder BA, Darby G, Richman DD. HIV with reduced sensitivity to zidovudine (AZT) isolated during prolonged therapy. Science 1989; 243:1731.
  2. Hirsch MS, Günthard HF, Schapiro JM, et al. Antiretroviral drug resistance testing in adult HIV-1 infection: 2008 recommendations of an International AIDS Society-USA panel. Clin Infect Dis 2008; 47:266.
  3. Japour AJ, Mayers DL, Johnson VA, et al. Standardized peripheral blood mononuclear cell culture assay for determination of drug susceptibilities of clinical human immunodeficiency virus type 1 isolates. The RV-43 Study Group, the AIDS Clinical Trials Group Virology Committee Resistance Working Group. Antimicrob Agents Chemother 1993; 37:1095.
  4. Kellam P, Larder BA. Recombinant virus assay: a rapid, phenotypic assay for assessment of drug susceptibility of human immunodeficiency virus type 1 isolates. Antimicrob Agents Chemother 1994; 38:23.
  5. Abstracts of the 3rd International Workshop on Adverse Drug Reactions and Lipodystrophy in HIV. 23-26 October 2001. Athens, Greece. Antivir Ther 2001; 6 Suppl 4:3.
  6. Wrin T, Huang W, Yap J, et al. Evaluating HIV-1 co-receptor usage and inhibitors of virus entry using recombinant virus assays. Antivir Ther 2001; 6:3.
  7. Limoli K, Huang W, Toma S, et al. Validation and performance characteristics of the PhenoSense HIV fusion inhibitor susceptibility assay. 45th Annual ICAAC, Washington DC, 2005. Abstract H-1076.
  8. Hertogs K, de Béthune MP, Miller V, et al. A rapid method for simultaneous detection of phenotypic resistance to inhibitors of protease and reverse transcriptase in recombinant human immunodeficiency virus type 1 isolates from patients treated with antiretroviral drugs. Antimicrob Agents Chemother 1998; 42:269.
  9. Petropoulos CJ, Parkin NT, Limoli KL, et al. A novel phenotypic drug susceptibility assay for human immunodeficiency virus type 1. Antimicrob Agents Chemother 2000; 44:920.
  10. Ross L, Boulmé R, Fisher R, et al. A direct comparison of drug susceptibility to HIV type 1 from antiretroviral experienced subjects as assessed by the antivirogram and PhenoSense assays and by seven resistance algorithms. AIDS Res Hum Retroviruses 2005; 21:933.
  11. Zhang J, Rhee SY, Taylor J, Shafer RW. Comparison of the precision and sensitivity of the Antivirogram and PhenoSense HIV drug susceptibility assays. J Acquir Immune Defic Syndr 2005; 38:439.
  12. Kuritzkes DR. Preventing and managing antiretroviral drug resistance. AIDS Patient Care STDS 2004; 18:259.
  13. Daar ES. Antiretroviral resistance in clinical practice. J Int Assoc Physicians AIDS Care (Chic) 2003; 2 Suppl 1:S4.
  14. Underwood MR, Ross LL, Irlbeck DM, et al. Sensitivity of phenotypic susceptibility analyses for nonthymidine nucleoside analogues conferred by K65R or M184V in mixtures with wild-type HIV-1. J Infect Dis 2009; 199:84.
  15. Cunningham S, Ank B, Lewis D, et al. Performance of the applied biosystems ViroSeq human immunodeficiency virus type 1 (HIV-1) genotyping system for sequence-based analysis of HIV-1 in pediatric plasma samples. J Clin Microbiol 2001; 39:1254.
  16. Erali M, Page S, Reimer LG, Hillyard DR. Human immunodeficiency virus type 1 drug resistance testing: a comparison of three sequence-based methods. J Clin Microbiol 2001; 39:2157.
  17. Collin G, Descamps D, Telles F, et al. Differences in protease and reverse transcriptase sequences between the TruGene HIV-1 genotyping kit (Visible Genetics) and the ViroSeq genotyping system (PE Applied biosystems). Antivir Ther 2000; 5:53.
  18. Demeter LM, D'Aquila R, Weislow O, et al. Interlaboratory concordance of DNA sequence analysis to detect reverse transcriptase mutations in HIV-1 proviral DNA. ACTG Sequencing Working Group. AIDS Clinical Trials Group. J Virol Methods 1998; 75:93.
  19. Schuurman R, Demeter L, Reichelderfer P, et al. Worldwide evaluation of DNA sequencing approaches for identification of drug resistance mutations in the human immunodeficiency virus type 1 reverse transcriptase. J Clin Microbiol 1999; 37:2291.
  20. Robert J, Fontaine E, Lambert C, et al. Inter-person variability in interpreting sequencing results for HIV-1 resistance testing. Antivir Ther 2000; 5:54.
  21. Koch N, Tamalet C, Tivoli N, et al. Comparison of two commercial assays for the detection of insertion mutations of HIV-1 reverse transcriptase. J Clin Virol 2001; 21:153.
  22. Fox ZV, Geretti AM, Kjaer J, et al. The ability of four genotypic interpretation systems to predict virological response to ritonavir-boosted protease inhibitors. AIDS 2007; 21:2033.
  23. Metzner KJ, Giulieri SG, Knoepfel SA, et al. Minority quasispecies of drug-resistant HIV-1 that lead to early therapy failure in treatment-naive and -adherent patients. Clin Infect Dis 2009; 48:239.
  24. Simen BB, Simons JF, Hullsiek KH, et al. Low-abundance drug-resistant viral variants in chronically HIV-infected, antiretroviral treatment-naive patients significantly impact treatment outcomes. J Infect Dis 2009; 199:693.
  25. Bellosillo NA, Bacheler L, Villacian J. HIV drug resistance tests: an update on methods for calculating phenotypic fold change from a viral genotype. Clin Infect Dis 2009; 48:687; author reply 687.
  26. Vermeiren H, Van Craenenbroeck E, Alen P, et al. Prediction of HIV-1 drug susceptibility phenotype from the viral genotype using linear regression modeling. J Virol Methods 2007; 145:47.
  27. Demeter L, Haubrich R. International perspectives on antiretroviral resistance. Phenotypic and genotypic resistance assays: methodology, reliability, and interpretations. J Acquir Immune Defic Syndr 2001; 26 Suppl 1:S3.
  28. Flandre P, Chappey C, Marcelin AG, et al. Phenotypic susceptibility to didanosine is associated with antiviral activity in treatment-experienced patients with HIV-1 infection. J Infect Dis 2007; 195:392.
  29. Harrigan PR, Montaner JS, Wegner SA, et al. World-wide variation in HIV-1 phenotypic susceptibility in untreated individuals: biologically relevant values for resistance testing. AIDS 2001; 15:1671.
  30. Rhee SY, Fessel WJ, Liu TF, et al. Predictive value of HIV-1 genotypic resistance test interpretation algorithms. J Infect Dis 2009; 200:453.
  31. Frentz D, Boucher CA, Assel M, et al. Comparison of HIV-1 genotypic resistance test interpretation systems in predicting virological outcomes over time. PLoS One 2010; 5:e11505.
  32. De Luca A, Cingolani A, Di Giambenedetto S, et al. Variable prediction of antiretroviral treatment outcome by different systems for interpreting genotypic human immunodeficiency virus type 1 drug resistance. J Infect Dis 2003; 187:1934.
  33. Liu TF, Shafer RW. Web resources for HIV type 1 genotypic-resistance test interpretation. Clin Infect Dis 2006; 42:1608.
  34. Van Laethem K, De Luca A, Antinori A, et al. A genotypic drug resistance interpretation algorithm that significantly predicts therapy response in HIV-1-infected patients. Antivir Ther 2002; 7:123.
  35. Desai S, Kyriakides T, Holodniy M, et al. Evolution of genotypic resistance algorithms and their impact on the interpretation of clinical trials: an OPTIMA trial substudy. HIV Clin Trials 2007; 8:293.
  36. Shafer RW. Low-abundance drug-resistant HIV-1 variants: finding significance in an era of abundant diagnostic and therapeutic options. J Infect Dis 2009; 199:610.
  37. Halvas EK, Wiegand A, Boltz VF, et al. Low frequency nonnucleoside reverse-transcriptase inhibitor-resistant variants contribute to failure of efavirenz-containing regimens in treatment- experienced patients. J Infect Dis 2010; 201:672.
  38. Paredes R, Lalama CM, Ribaudo HJ, et al. Pre-existing minority drug-resistant HIV-1 variants, adherence, and risk of antiretroviral treatment failure. J Infect Dis 2010; 201:662.
  39. Kuritzkes DR, Lalama CM, Ribaudo HJ, et al. Preexisting resistance to nonnucleoside reverse-transcriptase inhibitors predicts virologic failure of an efavirenz-based regimen in treatment-naive HIV-1-infected subjects. J Infect Dis 2008; 197:867.
  40. Johnson JA, Li JF, Wei X, et al. Minority HIV-1 drug resistance mutations are present in antiretroviral treatment-naïve populations and associate with reduced treatment efficacy. PLoS Med 2008; 5:e158.
  41. Garcia-Diaz A, Lok CB, Madge S, et al. Detection of low-frequency K103N mutants after unstructured discontinuation of efavirenz in the presence of the CYP2B6 516 TT polymorphism. J Antimicrob Chemother 2008; 62:1188.
  42. Chabria SB, Gupta S, Kozal MJ. Deep sequencing of HIV: clinical and research applications. Annu Rev Genomics Hum Genet 2014; 15:295.
  43. Lataillade M, Chiarella J, Yang R, et al. Prevalence and clinical significance of HIV drug resistance mutations by ultra-deep sequencing in antiretroviral-naïve subjects in the CASTLE study. PLoS One 2010; 5:e10952.
  44. Le T, Chiarella J, Simen BB, et al. Low-abundance HIV drug-resistant viral variants in treatment-experienced persons correlate with historical antiretroviral use. PLoS One 2009; 4:e6079.
  45. Hammer SM, Eron JJ Jr, Reiss P, et al. Antiretroviral treatment of adult HIV infection: 2008 recommendations of the International AIDS Society-USA panel. JAMA 2008; 300:555.
  46. Moyle GJ, Wildfire A, Mandalia S, et al. Epidemiology and predictive factors for chemokine receptor use in HIV-1 infection. J Infect Dis 2005; 191:866.
  47. Brumme ZL, Goodrich J, Mayer HB, et al. Molecular and clinical epidemiology of CXCR4-using HIV-1 in a large population of antiretroviral-naive individuals. J Infect Dis 2005; 192:466.
  48. Wilkin TJ, Su Z, Kuritzkes DR, et al. HIV type 1 chemokine coreceptor use among antiretroviral-experienced patients screened for a clinical trial of a CCR5 inhibitor: AIDS Clinical Trial Group A5211. Clin Infect Dis 2007; 44:591.
  49. Coakley E. HIV tropism testing in the clinic. J Viral Entry 2007; 3:10.
  50. Koot M, Keet IP, Vos AH, et al. Prognostic value of HIV-1 syncytium-inducing phenotype for rate of CD4+ cell depletion and progression to AIDS. Ann Intern Med 1993; 118:681.
  51. Panel on Antiretroviral Guidelines for Adults and Adolescents. Guidelines for the use of antiretroviral agents in HIV-1-infected adults and adolescents. Department of Health and Human Services. Available at http://aidsinfo.nih.gov/contentfiles/lvguidelines/AdultandAdolescentGL.pdf (Accessed on May 01, 2014).
  52. Hosoya N, Su Z, Wilkin T, et al. Assessing human immunodeficiency virus type 1 tropism: Comparison of assays using replication-competent virus versus plasma-derived pseudotyped virions. J Clin Microbiol 2009; 47:2604.
  53. Lin NH, Kuritzkes DR. Tropism testing in the clinical management of HIV-1 infection. Curr Opin HIV AIDS 2009; 4:481.
  54. Whitcomb JM, Huang W, Fransen S, et al. Development and characterization of a novel single-cycle recombinant-virus assay to determine human immunodeficiency virus type 1 coreceptor tropism. Antimicrob Agents Chemother 2007; 51:566.
  55. Reeves JD, Coakley E, Petropoulos CJ, Whitcomb JM. An enhanced sensitivity Trofile HIV coreceptor tropism assay for selecting patients for therapy with entry inhibitors targeting CCR5: a review of analytical and clinical studies. J Viral Entry 2009; 3:94.
  56. Raymond S, Delobel P, Mavigner M, et al. Correlation between genotypic predictions based on V3 sequences and phenotypic determination of HIV-1 tropism. AIDS 2008; 22:F11.
  57. Coakley E, Reeves JD, Huang W, et al. Comparison of human immunodeficiency virus type 1 tropism profiles in clinical samples by the Trofile and MT-2 assays. Antimicrob Agents Chemother 2009; 53:4686.
  58. Su Z, Gulick RM, Krambrink A, et al. Response to vicriviroc in treatment-experienced subjects, as determined by an enhanced-sensitivity coreceptor tropism assay: reanalysis of AIDS clinical trials group A5211. J Infect Dis 2009; 200:1724.
  59. Trouplin V, Salvatori F, Cappello F, et al. Determination of coreceptor usage of human immunodeficiency virus type 1 from patient plasma samples by using a recombinant phenotypic assay. J Virol 2001; 75:251.
  60. Kagan RM, Johnson EP, Siaw M, et al. A genotypic test for HIV-1 tropism combining Sanger sequencing with ultradeep sequencing predicts virologic response in treatment-experienced patients. PLoS One 2012; 7:e46334.
  61. McGovern RA, Thielen A, Mo T, et al. Population-based V3 genotypic tropism assay: a retrospective analysis using screening samples from the A4001029 and MOTIVATE studies. AIDS 2010; 24:2517.
  62. McGovern RA, Thielen A, Portsmouth S, et al. Population-based sequencing of the V3-loop can predict the virological response to maraviroc in treatment-naive patients of the MERIT trial. J Acquir Immune Defic Syndr 2012; 61:279.
  63. Swenson LC, Mo T, Dong WW, et al. Deep sequencing to infer HIV-1 co-receptor usage: application to three clinical trials of maraviroc in treatment-experienced patients. J Infect Dis 2011; 203:237.
  64. Swenson LC, Mo T, Dong WW, et al. Deep V3 sequencing for HIV type 1 tropism in treatment-naive patients: a reanalysis of the MERIT trial of maraviroc. Clin Infect Dis 2011; 53:732.