title: Comparison of capillary electrophoresis sequencing with the new CEQ 2000 DNA Analysis System to conventional gel based systems for HIV drug resistance analysis.
abstract: To date the majority of sequencing technologies have been based on use of gel plates. In this study sequencing by capillary electrophoresis for HIV-1 genotyping on the CEQ 2000 sequencer (Beckman Coulter Inc.) has been investigated and compared to an 'in house' protocol on the Prism-377 sequencer (Applied Biosystems) and to the HIV-1 TruGene kit (Visible Genetics Inc.), two gel plate-based systems. Plasma from 20 HAART-treated patients with virological failure were analyzed for protease (PR) and reverse transcriptase (RT) genes. A total of 520 RT codons (26/patient) and 360 PR codons (18/patient) related to antiretroviral drug resistance were evaluated. The overall agreement between CEQ 2000 and Prism-377 results was 100% for the RT and PR primary and secondary mutations. The overall agreement between CEQ 2000 and TruGene was 100% for primary and > or =97% for secondary mutations. Discrepant results would have never led to errors in genotype interpretation. Performances for a 24 patients/week/one technician genotyping throughput were analyzed. For Prism-377, TruGene and CEQ 2000, manual processing required 5, 4 and 2,5 days, sequence data analysis needed additional 3, 1 and 2 days and cost/patient was approximately 49, 214 and 39 $, respectively. The CEQ 2000 sequencer offers a reliable alternative for fast and cost effective HIV drug resistance analysis.
authors.affiliation: Laboratoire de Virologie, Centre Hospitalier Régional et Université Victor Ségalen, Bordeaux, France. patrick.merel@chu-bordeaux.fr
authors.forename: P
authors.initials: P
authors.lastname: Merel
authors.forename: I
authors.initials: I
authors.lastname: Pellegrin
authors.forename: I
authors.initials: I
authors.lastname: Garrigue
authors.forename: A
authors.initials: A
authors.lastname: Caumont
authors.forename: M H
authors.initials: MH
authors.lastname: Schrive
authors.forename: V
authors.initials: V
authors.lastname: Birac
authors.forename: P
authors.initials: P
authors.lastname: Bonot
authors.forename: H
authors.initials: H
authors.lastname: Fleury
mesh_terms: D019380:Anti-HIV Agents / Q000494:pharmacology / Q000627:therapeutic use; D024882:Drug Resistance, Viral / Q000235:genetics*; D019075:Electrophoresis, Capillary / Q000295:instrumentation; D005838:Genotype; D015658:HIV Infections / Q000188:drug therapy / Q000821:virology; D016333:HIV Protease / Q000235:genetics; D054303:HIV Reverse Transcriptase / Q000235:genetics; D015497:HIV-1 / Q000187:drug effects / Q000235:genetics*; D006801:Humans; D011933:Reagent Kits, Diagnostic; D017422:Sequence Analysis, DNA / Q000295:instrumentation
chemical_list: D019380:Anti-HIV Agents; D011933:Reagent Kits, Diagnostic; D054303:HIV Reverse Transcriptase; D016333:HIV Protease