Concordance between allele-specific PCR and ultra-deep pyrosequencing for the detection of HIV-1 non-nucleoside reverse transcriptase inhibitor resistance mutations

Gillian M. Hunt, Lynn Morris, Anitha Moorthy, Ashraf Coovadia, Elaine J. Abrams, Renate Strehlau, Louise Kuhn, Deborah Persaud

Research output: Contribution to journalArticlepeer-review

Abstract

Recent advances in genotyping technologies have allowed for detection of HIV-1 drug resistance mutations present at low levels. The presence and percentage of Y181C and K103N drug-resistant variants in the blood of 105 subtype C HIV-infected infants who failed single-dose nevirapine prophylaxis for HIV transmission were compared using two highly sensitive genotyping methods, allele-specific PCR (AS-PCR) and ultra-deep pyrosequencing. Significant correlations in detection between both methods were found for both Y181C (correlation coefficients of 0.94 [95% CI 0.91-0.96]) and K103N (0.89 [95% CI 0.84-0.92]) mutations. The majority of discordant specimens (3/5 Y181C and 8/11 K103N) had wild-type variants when population sequencing was used, but mutant variants were detectable at very low levels (≤5%) with either assay. This difference is most likely due to stochastic variations in the appearance of mutant variants. Overall, both AS-PCR and ultra-deep pyrosequencing methods have proven to be sensitive and accurate, and may confidently be used where feasible.

Original languageEnglish (US)
Pages (from-to)182-187
Number of pages6
JournalJournal of Virological Methods
Volume207
DOIs
StatePublished - Oct 2014

Keywords

  • Allele-specific PCR
  • HIV-1 drug resistance
  • Low-frequency NNRTI variants
  • Ultra-deep sequencing

ASJC Scopus subject areas

  • Virology

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