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Journal of Clinical Microbiology, August 2004, p. 3670-3674, Vol. 42, No. 8
0095-1137/04/$08.00+0     DOI: 10.1128/JCM.42.8.3670-3674.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Comparison of Oligonucleotide Ligation Assay and Consensus Sequencing for Detection of Drug-Resistant Mutants of Human Immunodeficiency Virus Type 1 in Peripheral Blood Mononuclear Cells and Plasma

Giovanina M. Ellis,1,{dagger} Madhumita Mahalanabis,1,{dagger},{ddagger} Ingrid A. Beck,1 Gregory Pepper,2 Amy Wright,2 Shannon Hamilton,2 Sarah Holte,3 Willscott E. Naugler,1,§ Diane M. Pawluk,1 Chung-Chen Li,1 and Lisa M. Frenkel1,2*

Department of Pediatrics,1 Laboratory Medicine, University of Washington,2 Fred Hutchinson Cancer Research Center, Seattle, Washington3

Received 20 March 2003/ Returned for modification 24 September 2003/ Accepted 24 April 2004

Drug-resistant mutants of human immunodeficiency virus type 1 (HIV-1) recede below the limit of detection of most assays applied to plasma when selective pressure is altered due to changes in antiretroviral treatment (ART). Viral variants with different mutations are selected by the new ART when replication is not suppressed or wild-type variants with greater replication fitness outgrow mutants following the cessation of ART. Mutants selected by past ART appear to persist in reservoirs even when not detected in the plasma, and when conferring cross-resistance they can compromise the efficacy of novel ART. Oligonucleotide ligation assay (OLA) of virus in plasma and peripheral blood mononuclear cells (PBMC) was compared to consensus sequence dideoxynucleotide chain terminator sequencing for detection of 91 drug resistance mutations that had receded below the limit of detection by sequencing of plasma. OLA of plasma virus detected 27.5% (95% confidence interval [CI], 19 to 39%) of mutant genotypes; consensus sequencing of the PBMC amplicon from the same specimen detected 23.1% (95% CI, 14 to 34%); and OLA of PBMC detected 53.8% (95% CI, 44 to 64%). These data suggest that concentrations of drug-resistant mutants were greater in PBMC than in plasma after changes in ART and indicate that the OLA was more sensitive than consensus sequencing in detecting low levels of select drug-resistant mutants.

* Corresponding author. Mailing address: 307 Westlake Ave. N, Suite 300, Seattle, WA 98109. Phone: (206) 987-5140. Fax: (206) 987-7311. E-mail: lfrenkel{at}u.washington.edu.

{dagger} Madhumita Mahalanabis and Giovanina Ellis contributed equally to this work.

{ddagger} Present address: Seattle Biomedical Research Institute and Department of Microbiology, University of Washington, Seattle, Wash.

§ Present address: Department of Medicine, University of California in San Diego, San Diego, Calif.

Journal of Clinical Microbiology, August 2004, p. 3670-3674, Vol. 42, No. 8
0095-1137/04/$08.00+0     DOI: 10.1128/JCM.42.8.3670-3674.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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