Rapid molecular detection of the H275Y oseltamivir resistance gene mutation in circulating influenza A (H1N1) viruses.
In early 2008, drug susceptibility surveillance of influenza viruses in Europe revealed that some influenza A viruses (subtype H1N1) circulating during the winter season of 2007 and 2008 were resistant to the neuraminidase inhibitor, oseltamivir. This resistance arises due to a histidine to tyrosine substitution in the neuraminidase active site (H275Y in N1 nomenclature). Current methods to detect this mutation involve an end-point reverse transcription polymerase chain reaction followed by nucleotide sequencing. While accurate, this approach has the limitation of being time-consuming, labour-intensive and expensive. Herein we describe a one-step allelic discrimination assay which rapidly (2h) detects this resistance mutation. The sensitivity of the assay was as low as 10 copies per reaction and is capable of detecting the antiviral resistance mutation in a mixture of wild type H275 and mutant H275Y targets.
Carr
Michael J
MJ
National Virus Reference Laboratory, University College Dublin, Belfield, Dublin 4, Ireland.
Sayre
Naomi
N
Duffy
Margaret
M
Connell
Jeff
J
Hall
William W
WW
D000483:Alleles; D019943:Amino Acid Substitution; D000998:Antiviral Agents / Q000494:pharmacology*; D024882:Drug Resistance, Viral / Q000235:genetics*; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype / Q000187:drug effects* / Q000235:genetics / Q000302:isolation & purification; D007251:Influenza, Human / Q000821:virology*; D008826:Microbial Sensitivity Tests; D009154:Mutation; D009439:Neuraminidase / Q000235:genetics; D053139:Oseltamivir / Q000494:pharmacology*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity; D013997:Time Factors
D000998:Antiviral Agents; D053139:Oseltamivir; D009439:Neuraminidase