Population analysis of oseltamivir-resistant variants for the rapid prediction of drug susceptibility by real-time reverse transcription polymerase chain reaction.

Abstract

This study investigated whether quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR), using specific probes composed of locked nucleic acids (LNA/qRT-PCR), designed to evaluate H1N1 pdm09 H275Y, H3N2 E119V and R292K variant populations, could replace a neuraminidase (NA) inhibition assay to determine the 50% inhibitory concentration (IC₅₀) of NA activity.For H1N1 pdm09, when the H275Y variant RNA load was 50% or 70% and the infective H275Y variant load was 40% or 70%, the IC₅₀ were >10- and 100-fold higher, respectively, than that of the wild-type (WT) strain. For H3N2, when the E119V RNA load and infective E119V variant load were >90% and >60%, respectively, the IC₅₀ of the mixed sample was >10-fold higher than that of the WT strain. The variant-mixed samples with a 70% or 80% R292K variant RNA load and a 60% or 70% infective R292K variant load exhibited >10- and 100-fold decreased susceptibility, respectively, compared with that of the WT. A positive correlation between the variant RNA load and infective variant load populations was observed.The LNA/qRT-PCR method can be used to improve the treatment and management of patients during antiviral therapy for influenza virus infection.