Comparison of illumina and 454 deep sequencing in participants failing raltegravir-based antiretroviral therapy

Jonathan Z. Li, Brad Chapman, Patrick Charlebois, Oliver Hofmann, Brian Weiner, Alyssa J. Porter, Reshmi Samuel, Saran Vardhanabhuti, Lu Zheng, Joseph Eron, Babafemi Taiwo, Michael C. Zody, Matthew R. Henn, Daniel R. Kuritzkes, Winston Hide, Roy M. Matining, Cara C. Wilson, Baiba I. Berzins, Edward P. Acosta, Barbara BastowPeter S. Kim, Sarah W. Read, Jennifer Janik, Debra S. Meres, Michael M. Lederman, Lori Mong-Kryspin, Karl E. Shaw, Louis G. Zimmerman, Randi Leavitt, Guy De La Rosa, Amy Jennings

Research output: Contribution to journalArticle

24 Scopus citations

Abstract

Background: The impact of raltegravir-resistant HIV-1 minority variants (MVs) on raltegravir treatment failure is unknown. Illumina sequencing offers greater throughput than 454, but sequence analysis tools for viral sequencing are needed. We evaluated Illumina and 454 for the detection of HIV-1 raltegravir-resistant MVs. Methods: A5262 was a single-arm study of raltegravir and darunavir/ritonavir in treatment-naïve patients. Pre-treatment plasma was obtained from 5 participants with raltegravir resistance at the time of virologic failure. A control library was created by pooling integrase clones at predefined proportions. Multiplexed sequencing was performed with Illumina and 454 platforms at comparable costs. Illumina sequence analysis was performed with the novel snp-assess tool and 454 sequencing was analyzed with V-Phaser. Results: Illumina sequencing resulted in significantly higher sequence coverage and a 0.095% limit of detection. Illumina accurately detected all MVs in the control library at ≥0.5% and 7/10 MVs expected at 0.1%. 454 sequencing failed to detect any MVs at 0.1% with 5 false positive calls. For MVs detected in the patient samples by both 454 and Illumina, the correlation in the detected variant frequencies was high (R2 = 0.92, P<0.001). Illumina sequencing detected 2.4-fold greater nucleotide MVs and 2.9-fold greater amino acid MVs compared to 454. The only raltegravir-resistant MV detected was an E138K mutation in one participant by Illumina sequencing, but not by 454. Conclusions: In participants of A5262 with raltegravir resistance at virologic failure, baseline raltegravir-resistant MVs were rarely detected. At comparable costs to 454 sequencing, Illumina demonstrated greater depth of coverage, increased sensitivity for detecting HIV MVs, and fewer false positive variant calls.

Original languageEnglish (US)
Article numbere90485
JournalPloS one
Volume9
Issue number3
DOIs
StatePublished - Mar 6 2014

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)
  • Agricultural and Biological Sciences(all)
  • General

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    Li, J. Z., Chapman, B., Charlebois, P., Hofmann, O., Weiner, B., Porter, A. J., Samuel, R., Vardhanabhuti, S., Zheng, L., Eron, J., Taiwo, B., Zody, M. C., Henn, M. R., Kuritzkes, D. R., Hide, W., Matining, R. M., Wilson, C. C., Berzins, B. I., Acosta, E. P., ... Jennings, A. (2014). Comparison of illumina and 454 deep sequencing in participants failing raltegravir-based antiretroviral therapy. PloS one, 9(3), [e90485]. https://doi.org/10.1371/journal.pone.0090485