Mullins Molecular Retrovirology Lab

  • Department of Microbiology
  • School of Medicine
  • University of Washington
University of Washington/Fred Hutch Center for AIDS Research

Citation Information

Ko, D., McLaughlin, S., Deng, W., Mullins, J.I., Dragavon, J., Harb, S., Coombs, R.W., Frenkel, L.M.(2024).Development and validation of a genotypic assay to quantify CXCR4- and CCR5-tropic human immunodeficiency virus type-1 (HIV-1) populations and a comparison to Trofile™ Viruses 2024 16, 510(pubmed) (doi)


HIV-1 typically infects cells via the CD4 receptor and CCR5 or CXCR4 co-receptors. Maraviroc is a CCR5-specific viral entry inhibitor; knowledge of viral co-receptor specificity is important prior to usage. We developed and validated an economical V3-env Illumina-based assay to detect and quantify the frequency of viruses utilizing each co-receptor. Plasma from 54 HIV+ participants (subtype B) was tested. The viral template cDNA was generated from plasma RNA with unique molecular identifiers (UMIs). The sequences were aligned and collapsed by the UMIs with a custom bioinformatics pipeline. Co-receptor usage, determined by codon analysis and online phenotype predictors PSSM and Geno2pheno, were compared to existing Trofile™ data. The cost of V3-UMI was tallied. The sequences interpreted by Geno2pheno using the most conservative cut-off, a 2% false-positive-rate (FPR), predicted CXCR4 usage with the greatest sensitivity (76%) and specificity (100%); PSSM and codon analysis had similar sensitivity and lower specificity. Discordant Trofile™ and genotypic results were more common when participants had specimens from different dates analyzed by either assay. V3-UMI reagents cost USD$62/specimen. A batch of <=20 specimens required 5 h of technical time across 1.5 days. V3-UMI predicts HIV tropism at a sensitivity and specificity similar to those of Trofile™, is relatively inexpensive, and could be performed by most central laboratories. The adoption of V3-UMI could expand HIV drug therapeutic options in lower-resource settings that currently do not have access to phenotypic HIV tropism testing.