title: Limited antigenic diversity of Plasmodium falciparum apical membrane antigen 1 supports the development of effective multi-allele vaccines.
abstract: BACKGROUND
  Polymorphism in antigens is a common mechanism for immune evasion used by many important pathogens, and presents major challenges in vaccine development. In malaria, many key immune targets and vaccine candidates show substantial polymorphism. However, knowledge on antigenic diversity of key antigens, the impact of polymorphism on potential vaccine escape, and how sequence polymorphism relates to antigenic differences is very limited, yet crucial for vaccine development. Plasmodium falciparum apical membrane antigen 1 (AMA1) is an important target of naturally-acquired antibodies in malaria immunity and a leading vaccine candidate. However, AMA1 has extensive allelic diversity with more than 60 polymorphic amino acid residues and more than 200 haplotypes in a single population. Therefore, AMA1 serves as an excellent model to assess antigenic diversity in malaria vaccine antigens and the feasibility of multi-allele vaccine approaches. While most previous research has focused on sequence diversity and antibody responses in laboratory animals, little has been done on the cross-reactivity of human antibodies.
  
  
  METHODS
  We aimed to determine the extent of antigenic diversity of AMA1, defined by reactivity with human antibodies, and to aid the identification of specific alleles for potential inclusion in a multi-allele vaccine. We developed an approach using a multiple-antigen-competition enzyme-linked immunosorbent assay (ELISA) to examine cross-reactivity of naturally-acquired antibodies in Papua New Guinea and Kenya, and related this to differences in AMA1 sequence.
  
  
  RESULTS
  We found that adults had greater cross-reactivity of antibodies than children, although the patterns of cross-reactivity to alleles were the same. Patterns of antibody cross-reactivity were very similar between populations (Papua New Guinea and Kenya), and over time. Further, our results show that antigenic diversity of AMA1 alleles is surprisingly restricted, despite extensive sequence polymorphism. Our findings suggest that a combination of three different alleles, if selected appropriately, may be sufficient to cover the majority of antigenic diversity in polymorphic AMA1 antigens. Antigenic properties were not strongly related to existing haplotype groupings based on sequence analysis.
  
  
  CONCLUSIONS
  Antigenic diversity of AMA1 is limited and a vaccine including a small number of alleles might be sufficient for coverage against naturally-circulating strains, supporting a multi-allele approach for developing polymorphic antigens as malaria vaccines.
authors.affiliation: The Burnet Institute of Medical Research and Public Health, 85 Commercial Road, Melbourne, Victoria, 3004, Australia. terheggen@gmx.de.|Department of Medicine, University of Melbourne, Melbourne, Victoria, Australia. terheggen@gmx.de.
authors.forename: Ulrich
authors.initials: U
authors.lastname: Terheggen
authors.affiliation: The Burnet Institute of Medical Research and Public Health, 85 Commercial Road, Melbourne, Victoria, 3004, Australia. damiendrew@burnet.edu.au.
authors.forename: Damien R
authors.initials: DR
authors.lastname: Drew
authors.affiliation: Walter and Eliza Hall Institute, Melbourne, Australia. hodder@wehi.edu.au.
authors.forename: Anthony N
authors.initials: AN
authors.lastname: Hodder
authors.affiliation: The Burnet Institute of Medical Research and Public Health, 85 Commercial Road, Melbourne, Victoria, 3004, Australia. cross.nadia@gmail.com.
authors.forename: Nadia J
authors.initials: NJ
authors.lastname: Cross
authors.affiliation: Centre for Geographic Medicine, Coast, Kenya Medical Research Institute, Kilifi, Kenya. ckmugyenyi@gmail.com.
authors.forename: Cleopatra K
authors.initials: CK
authors.lastname: Mugyenyi
authors.affiliation: Walter and Eliza Hall Institute, Melbourne, Australia. barry@wehi.edu.au.|Department of Medical Biology, University of Melbourne, Melbourne, Victoria, Australia. barry@wehi.edu.au.
authors.forename: Alyssa E
authors.initials: AE
authors.lastname: Barry
authors.affiliation: La Trobe University, Melbourne, Australia. r.anders@latrobe.edu.au.
authors.forename: Robin F
authors.initials: RF
authors.lastname: Anders
authors.affiliation: Walter Reed Army Institute, Silver Spring, MD, USA. sheetij.dutta@us.army.mil.
authors.forename: Sheetij
authors.initials: S
authors.lastname: Dutta
authors.affiliation: Centre for Geographic Medicine, Coast, Kenya Medical Research Institute, Kilifi, Kenya. sheetij.dutta@us.army.mil.
authors.forename: Faith H A
authors.initials: FH
authors.lastname: Osier
authors.affiliation: The Burnet Institute of Medical Research and Public Health, 85 Commercial Road, Melbourne, Victoria, 3004, Australia. salennaelliott@yahoo.com.
authors.forename: Salenna R
authors.initials: SR
authors.lastname: Elliott
authors.affiliation: Papua New Guinea Institute of Medical Research, Madang, Papua New Guinea. nicolas.senn@gmail.com.|Swiss Tropical and Public Health Institute, Basel, Switzerland. nicolas.senn@gmail.com.
authors.forename: Nicolas
authors.initials: N
authors.lastname: Senn
authors.affiliation: Walter and Eliza Hall Institute, Melbourne, Australia. d.stanisic@griffith.edu.au.|Papua New Guinea Institute of Medical Research, Madang, Papua New Guinea. d.stanisic@griffith.edu.au.
authors.forename: Danielle I
authors.initials: DI
authors.lastname: Stanisic
authors.affiliation: Centre for Geographic Medicine, Coast, Kenya Medical Research Institute, Kilifi, Kenya. kmarsh@kemri-wellcome.org.
authors.forename: Kevin
authors.initials: K
authors.lastname: Marsh
authors.affiliation: Papua New Guinea Institute of Medical Research, Madang, Papua New Guinea. peter.siba@pngimr.org.pg.
authors.forename: Peter M
authors.initials: PM
authors.lastname: Siba
authors.affiliation: Walter and Eliza Hall Institute, Melbourne, Australia. mueller@wehi.edu.au.|Department of Medical Biology, University of Melbourne, Melbourne, Victoria, Australia. mueller@wehi.edu.au.|Papua New Guinea Institute of Medical Research, Madang, Papua New Guinea. mueller@wehi.edu.au.
authors.forename: Ivo
authors.initials: I
authors.lastname: Mueller
authors.affiliation: The Burnet Institute of Medical Research and Public Health, 85 Commercial Road, Melbourne, Victoria, 3004, Australia. Richards@burnet.edu.au.|Department of Medicine, University of Melbourne, Melbourne, Victoria, Australia. Richards@burnet.edu.au.|Department of Microbiology, Monash University, Clayton, Victoria, Australia. Richards@burnet.edu.au.
authors.forename: Jack S
authors.initials: JS
authors.lastname: Richards
authors.affiliation: The Burnet Institute of Medical Research and Public Health, 85 Commercial Road, Melbourne, Victoria, 3004, Australia. beeson@burnet.edu.au.|Department of Medicine, University of Melbourne, Melbourne, Victoria, Australia. beeson@burnet.edu.au.|Department of Microbiology, Monash University, Clayton, Victoria, Australia. beeson@burnet.edu.au.
authors.forename: James G
authors.initials: JG
authors.lastname: Beeson
mesh_terms: D000293:Adolescent; D000328:Adult; D000483:Alleles; D000913:Antibodies, Protozoan / Q000276:immunology; D000940:Antigenic Variation; D000953:Antigens, Protozoan / Q000276:immunology*; D002648:Child; D002675:Child, Preschool; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007630:Kenya; D017780:Malaria Vaccines / Q000235:genetics / Q000276:immunology*; D016778:Malaria, Falciparum / Q000517:prevention & control*; D008565:Membrane Proteins / Q000276:immunology*; D008875:Middle Aged; D010219:Papua New Guinea; D010963:Plasmodium falciparum / Q000235:genetics / Q000276:immunology*; D011110:Polymorphism, Genetic; D015800:Protozoan Proteins / Q000276:immunology*
chemical_list: D000913:Antibodies, Protozoan; D000953:Antigens, Protozoan; D017780:Malaria Vaccines; D008565:Membrane Proteins; D015800:Protozoan Proteins; C090077:apical membrane antigen I, Plasmodium