title: Pandemic H1N1 2009 influenza virus with the H275Y oseltamivir resistance neuraminidase mutation shows a small compromise in enzyme activity and viral fitness.
abstract: BACKGROUND
  Resistance to the neuraminidase inhibitor oseltamivir can be conferred by a well-characterized mutation in the neuraminidase gene, H275Y. In human H1N1 viruses that circulated in the first years of the 21st century, this mutation carried a fitness cost and resistant viruses were rare. During the 2007-08 influenza season, oseltamivir-resistant viruses of H1N1 phenotype emerged and predominated. March 2009 saw the emergence of a novel H1N1 influenza pandemic. We examined whether the H275Y mutation affected neuraminidase enzyme activity or replication of the pandemic influenza virus.
  
  
  METHODS
  Using reverse genetics we engineered the H275Y mutation into the neuraminidase of a 2009 pandemic H1N1 virus and assessed the ability of this enzyme to desialylate mono- and multivalent substrates. The growth kinetics of wild-type and mutant viruses were assessed in Madin-Darby canine kidney (MDCK) and fully differentiated human airway epithelial (HAE) cells.
  
  
  RESULTS
  The presence of H275Y was associated with a 1.3-fold decrease in the affinity of the neuraminidase for a monovalent substrate and a 4-fold compromise in desialylation of multivalent substrate. This was associated with a fitness cost to viral replication in vitro, which only became apparent during competitive replication in the mucus-rich HAE culture system.
  
  
  CONCLUSIONS
  The neuraminidase protein of pandemic influenza isolates tolerates the H275Y mutation and this mutation confers resistance to oseltamivir. However, unlike seasonal H1N1 viruses isolated since 2007, the mutation is not associated with any fitness advantage and thus is unlikely to predominate without further antigenic drift, compensating mutations or intense selection pressure.
authors.affiliation: Section of Virology, Faculty of Medicine, Wright Fleming Institute, Imperial College London, Norfolk Place, London W2 1PG, UK.
authors.forename: Daniel W
authors.initials: DW
authors.lastname: Brookes
authors.forename: Shahajahan
authors.initials: S
authors.lastname: Miah
authors.forename: Angie
authors.initials: A
authors.lastname: Lackenby
authors.forename: Lorian
authors.initials: L
authors.lastname: Hartgroves
authors.forename: Wendy S
authors.initials: WS
authors.lastname: Barclay
mesh_terms: D000818:Animals; D000998:Antiviral Agents / Q000494:pharmacology*; D002478:Cells, Cultured; D004285:Dogs; D024882:Drug Resistance, Viral; D004847:Epithelial Cells / Q000821:virology; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype / Q000201:enzymology / Q000235:genetics / Q000254:growth & development / Q000502:physiology*; D007251:Influenza, Human / Q000821:virology; D020125:Mutation, Missense; D009439:Neuraminidase / Q000235:genetics* / Q000378:metabolism; D053139:Oseltamivir / Q000494:pharmacology*; D012794:Sialic Acids / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism; D014779:Virus Replication
chemical_list: D000998:Antiviral Agents; D012794:Sialic Acids; D014764:Viral Proteins; D053139:Oseltamivir; C487630:NA protein, influenza A virus; D009439:Neuraminidase