Brief Report Limited Evolution of HIV Antiretroviral Drug Resistance-Associated Mutations During the Performance of Drug Resistance Testing *Chris Birch, *Tracey Middleton, †Gillian Hales, †David Cooper, †Matthew Law, ‡Suzanne Crowe, §Jennifer Hoy, and †Sean Emery *Victorian Infectious Diseases Reference Laboratory, North Melbourne; †National Centre for HIV Epidemiology and Clinical Research, University of New South Wales, Sydney; ‡Burnet Centre, Melbourne; and §Alfred Hospital and Department of Medicine, Monash University, Melbourne, Australia Summary: We investigated the evolution of HIV reverse transcriptase (RT)– and protease-associated antiretroviral (ARV) drug resistance mutations during the time taken to perform genotypic drug resistance testing. Thirty treatment-experienced pa- tients who were adherent to therapy and who underwent genotypic drug resistance testing provided blood samples at randomization and when reviewing the test results (baseline). Patients remained on their existing therapy between randomization and baseline. The predominant HIV strains in 10 patients (33%) either lost and/or gained primary RT inhibitor (RTI)– or protease inhibitor (PI)–associated resistance mutations during the testing period. Of the 9 patients with RT mutations, 2 lost, 5 gained, and 2 both lost and gained RTI resistance mutations. One patient gained a significant PI- associated resistance mutation on an existing PI-resistant background. The evolution that occurred in the RT may have altered the effectiveness of subsequent ARV therapy in some patients. Neither viral load at randomization, ARV drug class used at ran- domization, time between collection of blood samples, duration of current therapy, nor number of ARV drugs used influenced gain or loss of resistance mutations. There was a significant association between duration of previous ARV therapy and gain of RTI- associated resistance mutations (p  .02), however. In general, our results suggest that patients should continue current therapy until test results are available. A few patients would be expected to gain ARV drug-associated resistance mutations during this time, however. Key Words: Antiretroviral drug therapy—Drug resistance testing— Resistance mutations—Evolution in reverse transcriptase and protease. Antiretroviral (ARV) drug resistance testing has be- come the standard of care in many developed countries because of the potential for the test results to have an impact on subsequent virologic responses to new thera- pies. Several studies examining the utility of resistance testing have shown that a sustained reduction in HIV viral load can be achieved by modifying therapy in re- sponse to the results (1–4). Although it has been recom- mended that blood used for resistance testing be drawn before the current drug regimen is stopped (5), no guide- lines are available to suggest whether treatment should be continued or discontinued during the time testing is being done. Using conventional assays, many resistance mutations rapidly become undetectable after ceasing therapy (6,7). Minor populations expressing important resistance mu- The NCHECR is funded by the Commonwealth Department of Health and Aged Care. The following companies provided financial support: Diagnostic Technology, Perkin Elmer Biosystems, Abbott Australasia, Boehringer Ingelheim, Bristol-Myers Squibb, Glaxo- SmithKline, Merck Sharpe and Dohme, and Roche Pharmaceuticals. Address correspondence and reprint requests to Chris Birch, Victo- rian Infectious Diseases Reference Laboratory, 10 Wreckyn Street, North Melbourne 3051, Victoria, Australia; e-mail: chris.birch@ mh.org.au Manuscript received June 25, 2002; accepted October 21, 2002. JAIDS Journal of Acquired Immune Deficiency Syndromes 32:57–61 © 2003 Lippincott Williams & Wilkins, Inc., Philadelphia 57