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Open Access Highly Accessed Methodology

A genotypic HIV-1 proviral DNA coreceptor tropism assay: characterization in viremic subjects

Jennifer Brown1*, Harold Burger2, Barbara Weiser2, Richard B Pollard1, Xiao-Dong Li2, Lynell J Clancy1, Russell E Baumann3, Amy A Rogers3, Hasnah B Hamdan3, Rick L Pesano3 and Ron M Kagan3

Author Affiliations

1 Division of Infectious Diseases, Department of Internal Medicine, University of California, Davis Medical Center, 4150 V. Street, PSSB-G500, Sacramento, CA, USA

2 University of California, Davis, School of Medicine, Davis, CA, USA

3 Department of Infectious Diseases, Quest Diagnostics Nichols Institute, San Juan Capistrano, CA, USA

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AIDS Research and Therapy 2014, 11:14  doi:10.1186/1742-6405-11-14

Published: 21 May 2014

Abstract

Background

HIV-1 coreceptor tropism testing is used to evaluate eligibility for CCR5 antagonist therapy. However, HIV-1 RNA-based tests are not suitable for virologically suppressed patients, therefore the use of proviral DNA tropism testing has been investigated. We describe a novel proviral DNA-based genotypic tropism assay and compare its performance to that of a sensitive HIV-1 RNA-based genotypic test.

Methods

Tropism was determined using HIV-1 plasma RNA and proviral DNA from 42 paired samples from patients with plasma viral loads ≥1000 HIV-1 RNA copies/mL. Proviral DNA sample types included whole blood, separated peripheral blood mononuclear cells resuspended in phosphate-buffered saline and peripheral blood mononuclear cells resuspended in spun plasma. The HIV-1 envelope V3 region was PCR-amplified, sequenced in triplicate, and analyzed for tropism with the geno2pheno algorithm using a 10% false-positive rate (FPR).

Results

Amplicons were obtained from proviral DNA and plasma RNA in 41/42 samples. Tropism predictions were highly concordant (93%–98%) between proviral DNA and plasma RNA, regardless of the proviral DNA isolation method. Non-R5 proviral DNA results were obtained for 100% of patients with detectable non-R5 plasma HIV-1 RNA results. Geno2pheno FPRs for proviral DNA and plasma RNA were highly correlated (Spearman rho = 0.86).

Conclusions

Our findings demonstrate that proviral DNA tropism determinations from whole blood or peripheral blood mononuclear cells were highly concordant with plasma HIV-1 RNA tropism determinations. This assay may be useful for screening virologically suppressed patients for CCR5-antagonist eligibility and for research purposes.

Keywords:
HIV-1 diagnostic tests; HIV-1 tropism; HIV-1 proviral tropism