title: A diagnostic HIV-1 tropism system based on sequence relatedness.
abstract: Key clinical studies for HIV coreceptor antagonists have used the phenotyping-based Trofile test. Meanwhile various simpler-to-do genotypic tests have become available that are compatible with standard laboratory equipment and Web-based interpretation tools. However, these systems typically analyze only the most prominent virus sequence in a specimen. We present a new diagnostic HIV tropism test not needing DNA sequencing. The system, XTrack, uses physical properties of DNA duplexes after hybridization of single-stranded HIV-1 env V3 loop probes to the clinical specimen. Resulting "heteroduplexes" possess unique properties driven by sequence relatedness to the reference and resulting in a discrete electrophoretic mobility. A detailed optimization process identified diagnostic probe candidates relating best to a large number of HIV-1 sequences with known tropism. From over 500 V3 sequences representing all main HIV-1 subtypes (Los Alamos database), we obtained a small set of probes to determine the tropism in clinical samples. We found a high concordance with the commercial TrofileES test (84.9%) and the Web-based tool Geno2Pheno (83.0%). Moreover, the new system reveals mixed virus populations, and it was successful on specimens with low virus loads or on provirus from leukocytes. A replicative phenotyping system was used for validation. Our data show that the XTrack test is favorably suitable for routine diagnostics. It detects and dissects mixed virus populations and viral minorities; samples with viral loads (VL) of <200 copies/ml are successfully analyzed. We further expect that the principles of the platform can be adapted also to other sequence-divergent pathogens, such as hepatitis B and C viruses.
authors.affiliation: Molecular Virology, Department of Biomedicine, University of Basel, Basel, Switzerland.
authors.forename: Suzanne
authors.initials: S
authors.lastname: Edwards
authors.affiliation: Molecular Virology, Department of Biomedicine, University of Basel, Basel, Switzerland.
authors.forename: Heinz
authors.initials: H
authors.lastname: Stucki
authors.affiliation: Molecular Virology, Department of Biomedicine, University of Basel, Basel, Switzerland.
authors.forename: Joëlle
authors.initials: J
authors.lastname: Bader
authors.affiliation: InPheno AG, Basel, Switzerland.
authors.forename: Vincent
authors.initials: V
authors.lastname: Vidal
authors.affiliation: Institute of Virology, University of Cologne, Cologne, Germany.
authors.forename: Rolf
authors.initials: R
authors.lastname: Kaiser
authors.affiliation: Division of Infectious Diseases and Hospital Epidemiology, University Hospital Basel, Basel, Switzerland.
authors.forename: Manuel
authors.initials: M
authors.lastname: Battegay
authors.affiliation: Molecular Virology, Department of Biomedicine, University of Basel, Basel, Switzerland thomas.klimkait@unibas.ch.
authors.forename: Thomas
authors.initials: T
authors.lastname: Klimkait
mesh_terms: D015331:Cohort Studies; D024202:Electrophoretic Mobility Shift Assay; D060005:Genotyping Techniques / Q000379:methods*; D015658:HIV Infections / Q000175:diagnosis* / Q000821:virology*; D015497:HIV-1 / Q000145:classification* / Q000235:genetics / Q000502:physiology*; D020180:Heteroduplex Analysis; D006801:Humans; D009693:Nucleic Acid Hybridization; D056189:Viral Tropism