title: Identification of amino acid substitutions supporting antigenic change of influenza A(H1N1)pdm09 viruses.
abstract: The majority of currently circulating influenza A(H1N1) viruses are antigenically similar to the virus that caused the 2009 influenza pandemic. However, antigenic variants are expected to emerge as population immunity increases. Amino acid substitutions in the hemagglutinin protein can result in escape from neutralizing antibodies, affect viral fitness, and change receptor preference. In this study, we constructed mutants with substitutions in the hemagglutinin of A/Netherlands/602/09 in an attenuated backbone to explore amino acid changes that may contribute to emergence of antigenic variants in the human population. Our analysis revealed that single substitutions affecting the loop that consists of amino acid positions 151 to 159 located adjacent to the receptor binding site caused escape from ferret and human antibodies elicited after primary A(H1N1)pdm09 virus infection. The majority of these substitutions resulted in similar or increased replication efficiency in vitro compared to that of the virus carrying the wild-type hemagglutinin and did not result in a change of receptor preference. However, none of the substitutions was sufficient for escape from the antibodies in sera from individuals that experienced both seasonal and pandemic A(H1N1) virus infections. These results suggest that antibodies directed against epitopes on seasonal A(H1N1) viruses contribute to neutralization of A(H1N1)pdm09 antigenic variants, thereby limiting the number of possible substitutions that could lead to escape from population immunity.
  
  
  OBJECTIVE
  Influenza A viruses can cause significant morbidity and mortality in humans. Amino acid substitutions in the hemagglutinin protein can result in escape from antibody-mediated neutralization. This allows the virus to reinfect individuals that have acquired immunity to previously circulating strains through infection or vaccination. To date, the vast majority of A(H1N1)pdm09 strains remain antigenically similar to the virus that caused the 2009 influenza pandemic. However, antigenic variants are expected to emerge as a result of increasing population immunity. We show that single amino acid substitutions near the receptor binding site were sufficient to escape from antibodies specific for A(H1N1)pdm09 viruses but not from antibodies elicited in response to infections with seasonal A(H1N1) and A(H1N1)pdm09 viruses. This study identified substitutions in A(H1N1)pdm09 viruses that support escape from population immunity but also suggested that the number of potential escape variants is limited by previous exposure to seasonal A(H1N1) viruses.
authors.affiliation: Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands.
authors.forename: Björn F
authors.initials: BF
authors.lastname: Koel
authors.affiliation: Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands Center for Pathogen Evolution, Department of Zoology, University of Cambridge, Cambridge, United Kingdom.
authors.forename: Ramona
authors.initials: R
authors.lastname: Mögling
authors.affiliation: Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands.
authors.forename: Salin
authors.initials: S
authors.lastname: Chutinimitkul
authors.affiliation: Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands Department of Pediatrics, Erasmus MC-Sophia, Rotterdam, The Netherlands.
authors.forename: Pieter L
authors.initials: PL
authors.lastname: Fraaij
authors.affiliation: Center for Pathogen Evolution, Department of Zoology, University of Cambridge, Cambridge, United Kingdom WHO Collaborating Centre for Modeling Evolution and Control of Emerging Infectious Diseases, University of Cambridge, Cambridge, United Kingdom.
authors.forename: David F
authors.initials: DF
authors.lastname: Burke
authors.affiliation: Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands.
authors.forename: Stefan
authors.initials: S
authors.lastname: van der Vliet
authors.affiliation: Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands.
authors.forename: Emmie
authors.initials: E
authors.lastname: de Wit
authors.affiliation: Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands.
authors.forename: Theo M
authors.initials: TM
authors.lastname: Bestebroer
authors.affiliation: Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands.
authors.forename: Guus F
authors.initials: GF
authors.lastname: Rimmelzwaan
authors.affiliation: Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands.
authors.forename: Albert D M E
authors.initials: AD
authors.lastname: Osterhaus
authors.affiliation: Center for Pathogen Evolution, Department of Zoology, University of Cambridge, Cambridge, United Kingdom WHO Collaborating Centre for Modeling Evolution and Control of Emerging Infectious Diseases, University of Cambridge, Cambridge, United Kingdom Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands.
authors.forename: Derek J
authors.initials: DJ
authors.lastname: Smith
authors.affiliation: Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands.
authors.forename: Ron A M
authors.initials: RA
authors.lastname: Fouchier
authors.affiliation: Department of Viroscience, Erasmus MC, Rotterdam, The Netherlands Center for Pathogen Evolution, Department of Zoology, University of Cambridge, Cambridge, United Kingdom m.degraaf@erasmusmc.nl.
authors.forename: Miranda
authors.initials: M
authors.lastname: de Graaf
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D057134:Antibodies, Neutralizing / Q000097:blood; D000914:Antibodies, Viral / Q000097:blood*; D000940:Antigenic Variation; D000956:Antigens, Viral / Q000235:genetics / Q000276:immunology*; D004252:DNA Mutational Analysis; D018985:Epitopes, B-Lymphocyte / Q000276:immunology; D005289:Ferrets; D040961:Genetic Drift; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000235:genetics / Q000276:immunology*; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype / Q000235:genetics / Q000276:immunology* / Q000502:physiology; D014779:Virus Replication
chemical_list: D057134:Antibodies, Neutralizing; D000914:Antibodies, Viral; D000956:Antigens, Viral; D018985:Epitopes, B-Lymphocyte; D019267:Hemagglutinin Glycoproteins, Influenza Virus