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Journal of Clinical Microbiology, February 2001, p. 454-459, Vol. 39, No. 2
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.2.454-459.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Comparison of DNA Sequencing and a Line Probe Assay for Detection of Human Immunodeficiency Virus Type 1 Drug Resistance Mutations in Patients Failing Highly Active Antiretroviral Therapy

Jean Servais,1,* Christine Lambert,1 Elodie Fontaine,1 Jean-Marc Plesséria,1 Isabelle Robert,1 Vic Arendt,1,2 Thérèse Staub,1,2 François Schneider,1,3 Robert Hemmer,1,2 Guy Burtonboy,4 and Jean-Claude Schmit1,2

Laboratoire de Rétrovirologie, Centre de Recherche Public-Santé,1 Service National des Maladies Infectieuses, Centre Hospitalier de Luxembourg,2 and Laboratoire National de Santé,3 Luxembourg, Luxembourg, and Unité de Virologie, Université Catholique de Louvain, Brussels, Belgium4

Received 18 July 2000/Returned for modification 23 October 2000/Accepted 7 November 2000

The resistance of human immunodeficiency virus type 1 (HIV-1) to drugs is a major cause of antiretroviral treatment failure. We have compared direct sequencing to a line probe assay (LiPA) for the detection of drug resistance-related mutations in 197 clinical samples, and we have investigated the sequential appearance of mutations under drug pressure. For 26 patients with virological failure despite the use of two nucleoside analogues and one protease inhibitor (indinavir [n = 6], ritonavir [n = 10], and saquinavir [n = 10]), genotypic resistance assays were carried out retrospectively every 3 months for up to 2 years by using direct sequencing (TruGene; Visible Genetics) and a LiPA for detection of mutations in the reverse transcriptase (INNO-LiPA HIV-1 RT; Innogenetics) and the protease (INNO-LiPA HIV Protease, prototype version; Innogenetics) genes. Comparison of the results from both assays found rare major discrepancies (<1% of codons analyzed). INNO-LiPA detected more wild-type-mutant mixtures than sequencing but suffered from a high rate of codon hybridization failures for the reverse transcriptase. LiPA detected earlier and more frequently than sequencing the transient mixed virus population that contained I84V, which appears before V82A in the protease sequence. Mutations M461, G48V, and L90M were often transient and drug pressure related. In conclusion, direct sequencing and LiPAs give concordant results for most clinical isolates. LiPAs are more sensitive for the detection of mixed virus populations. Mutation I84V appears in minor populations in the early steps of the pathways of resistance to indinavir and ritonavir. The fact that some mutations can be found only transiently and in minor virus populations highlights the importance of a low detection limit for resistance assays.

* Corresponding author. Mailing address: Laboratoire de Rétrovirologie, CRP-Santé, 4 rue E. Barblé, L-1210 Luxembourg, Luxembourg. Phone: 352-44116105. Fax: 352-44116113. E-mail: servais.j{at}retrovirology.lu.

Journal of Clinical Microbiology, February 2001, p. 454-459, Vol. 39, No. 2
0095-1137/01/$04.00+0   DOI: 10.1128/JCM.39.2.454-459.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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