Identification of cross-stage, cross-species malaria CD8 + T cell antigens

Data availability

The data that support the findings of this study are available in the Article and its Supplementary Information. The mass spectrometry proteomics data are available via ProteomeXchange with the identifier PXD077321. Source data are provided with this paper.

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Acknowledgements

We thank all patients and healthy volunteers; the clinic, laboratory and administrative staff; and field workers and study participants from Brazil, Mali and the USA. We also thank the Sequencing Facility of the MRC Human Immunology Unit at the University of Oxford for HLA genotyping, H. Liao for assistance with the NetMHC analysis and F. Salusto for providing the K562 HLA-I−/− cells and K562 HLA−/− cells overexpressing HLA-E*01:03. L.P.B.C., L.R.V.A., R.T.G. and C.J. are recipients of Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) fellowships; C.R.R.B. and B.C.M.-R. are fellows of the Fundação de Amparo à Pesquisa do Estado de Minas Gerais (FAPEMIG); and C.R.R.B., L.B.d.L. and G.C.M. are fellows of the Coordenação de Aperfeiçoamento de Pessoal de Ensino Superior (CAPES). The contributions of the NIH authors are considered works of the US government. The findings and conclusions presented in this paper are those of the authors and do not necessarily reflect the views of the NIH or the US Department of Health and Human Services.

Funding

This study was funded by the National Institutes of Health (NIH/NIAID R01 AI102063-01 to C.J. and B.W.; Amazonian ICEMR U19 AI089681 to C.J., L.R.V.A. and R.T.G.; NIH/NIAID U01AI165457 to B.W.; and P51OD011092 (support by the Office of the Director) to the Oregon National Primate Research Center); the National Institute of Science and Technology for Vaccines–CNPq (465293/2014-0 to C.J., D.B.P., L.R.V.A. and R.T.G.); CNPq (437851/2018-4 to C.J.); FAPEMIG (RED-00012-14, APQ-00653-16 and APQ-02962-18 to C.J.); the European Union’s Horizon 2020 research and innovation programme (733273 to A.V.S.H.); the Lemann Brazil Research Fund to C.J. and J.L.; a Silver Family Foundation–OHSU Faculty Excellence and Innovation Award to B.W.; the Fondazione per l’Istituto di Ricerca in Biomedicina to C.J.; and the Helmut Horten Foundation to C.J. This research was supported in part by the NIH Intramural Research Program.

Author information

Author notes

These authors contributed equally: Camila R. R. Barbosa, Luna B. de Lacerda, Paulo J. G. Bettencourt

Authors and Affiliations

Instituto René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Brazil

Camila R. R. Barbosa, Luna B. de Lacerda, Cristopher Gomes, Lídia P. B. Cordeiro, Guilherme C. Maia, Gregório G. Almeida, Beatriz C. Medeiros-Rodrigues, Camila M. Costa, Lis R. V. Antonelli, Ricardo T. Gazzinelli & Caroline Junqueira

Centro de Tecnologia em Vacinas, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil

Luna B. de Lacerda, Cristopher Gomes, Lídia P. B. Cordeiro, Gregório G. Almeida, Ricardo T. Gazzinelli & Caroline Junqueira

Jenner Institute, University of Oxford, Oxford, UK

Paulo J. G. Bettencourt, Annalisa Nicastri, Nicola Ternette & Adrian V. S. Hill

Center for Interdisciplinary Research in Health, Faculty of Medicine, Universidade Católica Portuguesa, Lisbon, Portugal

Paulo J. G. Bettencourt

Vaccine and Gene Therapy Institute, Oregon Health & Science University, Beaverton, OR, USA

David Morrow, Maya Aleshnick, Julie L. Mitchell, Thalia F. Hart, Derek Haumpy, Roxanne M. Gilbride, John B. Schell, Payton Kirtley, Scott G. Hansen, Klaus Früh & Brandon Wilder

Centro de Pesquisas em Medicina Tropical de Rondônia, Porto Velho, Brazil

Dhelio B. Pereira

Laboratory of Malaria Immunology and Vaccinology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, USA

Nicholas C. Poulton, John Woodford, Joel Goldberg & Patrick E. Duffy

Departmaneto de Bioquímica e Imunologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, Brazil

Cristopher Gomes, Guilherme C. Maia & Ricardo T. Gazzinelli

Institute for Research in Biomedicine, Università della Svizzera italiana, Bellinzona, Switzerland

Cristopher Gomes, Lídia P. B. Cordeiro, Marie Rose Schrimpf & Caroline Junqueira

Parasites and Microbes Research and Training Center, University of Sciences, Techniques and Technologies of Bamako, Bamako, Mali

Kadia Doumbia, Charles Arama & Moussa Niangaly

Max Planck Institute for Infection Biology, Berlin, Germany

Christina Ntalla, Moussa Niangaly & Silvia Portugal

Ragon Institute of Mass General, MIT and Harvard, Cambridge, MA, USA

Zezhou Zhao & Gaurav D. Gaiha

Program in Cellular and Molecular Medicine, Boston Children’s Hospital, Boston, MA, USA

Marie Rose Schrimpf & Judy Lieberman

Department of Pediatrics, Harvard Medical School, Boston, MA, USA

Marie Rose Schrimpf & Judy Lieberman

Chester Beatty Laboratories, Institute of Cancer Research, London, UK

Annalisa Nicastri

Program in Health Sciences and Technology, Harvard Medical School and Massachusetts Institute of Technology, Cambridge, MA, USA

Gaurav D. Gaiha

Division of Gastroenterology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA

Gaurav D. Gaiha

QIMR Berghofer, Brisbane, Queensland, Australia

John Woodford

School of Life Sciences, University of Dundee, Dundee, UK

Nicola Ternette

Authors

Camila R. R. Barbosa

Luna B. de Lacerda

Paulo J. G. Bettencourt

David Morrow

Dhelio B. Pereira

Maya Aleshnick

Julie L. Mitchell

Nicholas C. Poulton

Cristopher Gomes

Lídia P. B. Cordeiro

Kadia Doumbia

Christina Ntalla

Charles Arama

Zezhou Zhao

Guilherme C. Maia

Gregório G. Almeida

Marie Rose Schrimpf

Thalia F. Hart

Derek Haumpy

Beatriz C. Medeiros-Rodrigues

Camila M. Costa

Annalisa Nicastri

Roxanne M. Gilbride

John B. Schell

Payton Kirtley

Lis R. V. Antonelli

Gaurav D. Gaiha

Scott G. Hansen

Judy Lieberman

Ricardo T. Gazzinelli

Moussa Niangaly

John Woodford

Joel Goldberg

Klaus Früh

Silvia Portugal

Patrick E. Duffy

Nicola Ternette

Brandon Wilder

Adrian V. S. Hill

Caroline Junqueira

Contributions

C.J. conceived the study. C.J., N.T., A.V.S.H., B.W., P.E.D. and S.P. supervised this study. D.B.P. recruited the patients and healthy donors. J.G. and J.W. coordinated the CHMI study. C.R.R.B., L.B.d.L., P.J.G.B., L.R.V.A., G.D.G., S.G.H., J.L., R.T.G., M.N., J.G., K.F., S.P., P.E.D., N.T., B.W., A.V.S.H. and C.J. designed and discussed experiments. C.R.R.B., L.B.d.L., P.J.G.B., D.M., M.A., J.L.M., N.C.P., C.G., L.P.B.C., K.D., C.N., C.A., Z.Z., G.C.M., G.G.A., A.N., M.R.S., T.F.H., D.H., B.C.M.-R., C.M.C., A.N., R.M.G., J.B.S., P.K., N.T. and C.J. performed and analysed experiments. C.J., G.D.G., S.G.H., K.F., R.T.G., N.T., B.W. and A.V.S.H. contributed reagents. C.R.R.B., L.B.d.L., M.A., J.L.M., B.W. and C.J. prepared figures and helped with manuscript preparation.

Corresponding authors

Correspondence to

Brandon Wilder, Adrian V. S. Hill or Caroline Junqueira.

Ethics declarations

Competing interests

The authors declare no competing interests.

Peer review

Peer review information

Nature thanks Jake Baum, Opeyemi Oludada, Stephanie Yanow and the other, anonymous, reviewer(s) for their contribution to the peer review of this work. Peer reviewer reports are available.

Additional information

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Extended data figures and tables

Extended Data Fig. 1 Comparative analysis of the length distribution of Pv and human peptides and protein identity between Pv and Pf.

a, Distribution of peptide lengths (8–25 amino acids) for human and Pv peptides. b, Table of Pv proteins with the number of patients in which each protein was detected, and the percentage of sequence identity to Pf orthologs. c, Each point represents a parasite-peptide. The y axis indicates protein sequence identity to Pf, and the x axis shows the number of patients in which the peptide was detected. Colours denote patient frequency: pink (7 patients), orange (6), purple (5), blue (4), and green (3).

Extended Data Fig. 2 Antigen gene expression in humanized mouse livers infected with Pf or Pv.

a,b, Expression of genes corresponding to antigens identified by Pv immunopeptidomics during liver stage infection with Pf (a) or Pv (b). Parasite 18S rRNA was used as endogenous control. CSP and TRAP genes served as positive control. Data are shown as mean ± s.e.m. Statistical analysis was performed using one-way ANOVA with Tukey’s multiple comparisons test; exact P values are indicated.

Extended Data Fig. 3 Peptide validation by ex vivo IFNγ ELISpot.

a–e, PBMCs from a Brazilian cohort: Pv-infected (n = 24); Pf-infected (n = 7), endemic HD (n = 15); and non-endemic HD (n = 6). Cells were stimulated with peptides derived from a, 40S ribosomal proteins, b, 60S ribosomal protein, c, ETRAMPs, d, histone, and e, other proteins. f, PBMCs from a Malian cohort: Pf-infected individuals (n = 5) and endemic HD (n = 7). IFNγ responses are shown as SFC per 1 × 106 PBMCs. Each symbol represents one individual. Responses ≥30 spots are considered positive (red dashed line). Anti-CD3/anti-CD28 stimulation was used as positive control.

Extended Data Fig. 4 Pv peptides are presented by different HLA molecules.

a, HLA-ABC genotyping of all 7 patients in the immunopeptidome analysis. b–d, NetMHC bind analysis of peptides identified in each sample to their donor HLA type. b, Number of peptides identified as HLA-ABC binders for each sample. HLA-ABC genotyping of the donor of samples used in the immunopeptidomics assays. Alleles represented with same colour mean that they were shared across different samples. c, Total number of peptides predicted to bind to each sample. Bars represent the overall peptide counts for each individual. d, Length distribution of binder peptides. Histogram shows the frequency of peptides according to their amino-acid length, highlighting the predominance of 8–11-mer peptides typical of HLA class I binding.

Extended Data Fig. 5 Expression of tapasin and ERAP1 on reticulocytes.

a, Western blot of cell lysates from uninfected reticulocytes (uRetics) from three healthy donors, and cell lysates from Pv-infected reticulocytes (Pv iRetics) from three patients, were probed for the antigen-processing protein tapasin and ERAP1. PBMC was used as a positive control. Anti-β-actin and anti-F-actin were used as endogenous protein expression control for uninfected and infected cells, respectively. b, In vitro binding stability assay of all selected peptides pulsed in an HLA-A*02:01, HLA-E*01:01, and HLA-E*01:03 cell lines, including control peptide (SL9 for HLA-A*02:01 and VL9 for HLA-E*01:01 and HLA-E*01:03). Peptides marked with (*) whose mean MFI are higher than s.d. are considered to be binders. Peptides marked with (#) were in silico predicted to be HLA-A*02:01 binders.

Source data

Extended Data Fig. 6 Parasitaemia during Pk and Pcy infection of NHPs.

a, Parasitaemia of Pk as measured by qRT–PCR after each immunization. b, Antibodies to Pk CSP repeat region throughout immunization as measured by single dilution ELISA OD value. c, Parasitaemia of Pcy as measured by Giemsa-stained thin smears.

Extended Data Fig. 7 Peptide validation by ex vivo IFNγ ELISpot in a mouse model.

a–c, Splenocytes from Py-infected mice were isolated at day 12 after infection following intraperitoneal inoculation of 105 infected reticulocytes and stimulated with peptides derived from a, ETRAMPs, b, Histone and c, other proteins. Red symbols indicate infected mice (n = 6) and black symbols indicate uninfected controls (n = 3). Each symbol represents one mouse. IFNγ responses are shown as SFCs per 106 splenocytes. Responses ≥10 SFC are considered positive (#, red dashed line). Statistical analysis was performed using multiple unpaired Student’s t-tests; P values are indicated.

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Barbosa, C.R.R., de Lacerda, L.B., Bettencourt, P.J.G. et al. Identification of cross-stage, cross-species malaria CD8+ T cell antigens.

Nature (2026). https://doi.org/10.1038/s41586-026-10730-1

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Received: 16 May 2025

Accepted: 28 May 2026

Published: 01 July 2026

Version of record: 01 July 2026

DOI: https://doi.org/10.1038/s41586-026-10730-1