Long read technologies provides novel insight into the diversity of the MHC in Africa a roadmap to understanding HLA diversity in Africa
Martin Pollard Global Health and Populations – Human Genetics, Wellcome Sanger Institute Sandhu Group – Department of Medicine, University of Cambridge Introduction
• Africa - genetically most diverse region in the world • Hundreds of distinct populations • Evolutionary adaptation to selective pressures • Different causative loci between populations HLA
• MHC - one of the most genetically diverse regions • African HLA diversity high relative to Europeans • Understudied in Africa • 16835 classical sequences in IMGT 3.29.0.1 • Of those 575 sequences with African ethnic source tag Jan van Rood (1926-2017) MHC region: relevance
Immature Antigen Immature Antigen CD4+T cell CD8+ Antigen T cell Antigen Presenting Presenting Cell Cell • Infectious and non- TCR TCR MHCII MHCI infectious disease • Autoimmunity • Cancer + + CD4 CD8 • Vaccine response • Transplant medicine Mature helper Mature cytotoxic T cell T Cell (Th1 or Th2) (Tc)
Picture By user:Sjef - CC BY-SA 3.0 wikicommons Challenges in the study of the HLA region
• Genetic diversity – high heterozygosity • High levels of homopolymer repeats • Structural variation • Limited sequence data in reference resources (82.7% IMGT 3.29.0.1 sequences “partial”) • Lack of validation in African populations of current methods • Sanger based sequencing - resolution of phase • Primer based amplification • Capture Sequencing of the HLA region
MiSeq + Pacbio Pacbio sequencing N=(1706) (n=285)
Demultiplex
Consensus
A*01:01:01:01 B*07:02 A*01:01:01:01 C*33:10:01 B*07:02 C*33:10:01 Lab Process – Sample Management Lab Process PCR Lab Process QC & EM pool The workhorses Long Amplicon Analysis HLA Diversity in Africa panel
Population Samples Population Samples
Urban – Entebbe, Uganda 330 Morrocan Expat – Netherlands 25
Urban – Banfora, Burkina Faso 167 Igbo – Nigeria 25 Urban – Soweto, South Africa 204+196 Kalenjin – Kenya 25 Yoruba – Ibadan, Nigeria (YRI) 110 Ashanti – Ghana 25 Esan – Nigeria (ESN) 99 Zulu – South Africa 25 Maasai – Kinyawa, Kenya (MKK) 166 Egyptian – Egypt 17 Gambian – Western Division, 112 The Gambia (GWD) Fur (2 Borgu, 1 Mima, 1 18 Mende – Sierra Leone (MSL) 84 Karanga) – Sudan Burkinabe – Gouin, Burkina 25 Americans of African Ancestry, 62 Faso SW USA (ASW) Maasai – Kinyawa, Kenya (MKK) 25 Afro Caribbean – Barbados 79 (ACB) Mandinka – Gambia 25 Luhya – Webuye, Kenya (LWK) 97 Muganda – Uganda 25
Nama – South Africa 25
Phase 3 In progress 93 1,831 HLA typed persons IAVI Protocol C later this year 600 1,322 Full length full phase Phase 4 later this year 50 22 diverse populations A workflow for accurate HLA typing in Africans: A clinical and medical genetics resources
• Poor disambiguation of HLA types with Sanger sequencing • High concordance between SBT and PacBio sequencing (>95% at 2 digits) • PacBio sequencing more accurate when discordance observed • High levels of novelty Novel Allele Differences Novel Allele Genetic Origin Novel Allele Ethnic Origin Diversity of DQB1 HLA diversity in Africa in the global context
Alexander Mentzer Distribution of functionally important alleles
Gambian) • B*53:01:01 – Malaria Symptom Resistance Mende Afro Caribbean, Ashanti, Esan Burkina Faso, Esan, Ibo,
Ashanti Burkina Faso Ibo Luhya, Maasai, Yoruba Luhya Maasai Kalenjin
Yoruba Entebbe
Afro-Caribbean
B*53:01:01G Zulu Other allele Malaria endemic regions – World Bank/MARA/ARMA HLA panel as a resource for imputation
Alexander Mentzer and Alexander Dilthey African HLA panels as a resource for medical research • Does Human Variation influence vaccine response? • 2499 Samples • Multiplex Immunoassay against vaccine antigens (Total IgG) • Pertussis Toxin • Filamentous Haemagglutinin • Pertactin • Tetanus Toxin • Diphtheria Toxin • Haemophilus influenzae type b • Hepatitis B surface antigen • Measles virus
Alexander Mentzer HLA panel facilitates fine mapping of HLA signals
Alexander Mentzer Conclusion
• HLA diversity in Africa panel- largest HLA sequence to date • Informs clinical typing • Use of long range sequencing platforms • Novel alleles • Resolving ambiguity • A resource for medical genetics • Substantial improvement in imputation • Fine mapping • High levels of genetic diversity across Africa • Functional importance • Impact on clinical phenotypes • Availability • Managed access (EGA) • Imputation Server Acknowledgements
• Our many research participants from across Africa • Samples Alison Elliot (EMaB) • My co-author Alex Mentzer (University of Oxford) • • Brenna Henn and Eileen G. Hoal (Nama) • Wellcome Sanger Institute Manj Sandhu • Charles Rotimi (AADM) • Charles Agyemang and Karien Stronks (HELIUS) Deepti Gurdasani Cristina Pomilla • Endashaw Bekele (Amhara, Gumuz, Somali) Tarryn Porter • Fraser Pirie and Ayesha Motaya (DCC) Tommy Carstensen Mike Quail & the Sanger Seq R&D group • Hisham Hassan and Mihai Netea (Beja, Copts, Fulani, Fur, Mahas) Naomi Park • Pierre Zalloua (Sara, Toubou, Egyptians) PacBio Team • Richard Cooper (EpiTrans Ghana) • University of Oxford • Shabir Madhi (South African Infants GWAS) Adrian Hill Gill McVean • Sodiomon B. Sirma (Banfora VAC050, Gouin) Kathryn Auckland • NHGRI repository at Coriell Institute Alex Dilthey • IHWG • PacBio • Histogenetics
• Nicola Cahill IMGT/HLA • Swati Ranade • • John Harting • IAVI, USAID and Wellcome • John Baeten • Mark Thornber • Riki Aydeniz