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Host genetic factors can confer resistance against malaria, raising the question of whether this has led to evolutionary adaptation of parasite populations. In this study we investigated the correlation between host and parasite genetic variation in 4,171 Gambian and Kenya children ascertained with severe malaria due to Plasmodium falciparum. We identified a strong association between sickle haemoglobin (HbS) in the host and variation in three regions of the parasite genome, including nonsynonymous variants in the acyl-CoA synthetase family member PfACS8 on chromosome 2, in a second region of chromosome 2, and in a region containing structural variation on chromosome 11. The HbS-associated parasite alleles are in strong linkage disequilibrium and have frequencies which covary with the frequency of HbS across populations, in particular being much more common in Africa than other parts of the world. The estimated protective effect of HbS against severe malaria, as determined by comparison of cases with population controls, varies greatly according to the parasite genotype at these three loci. These findings open up a new avenue of enquiry into the biological and epidemiological significance of the HbS-associated polymorphisms in the parasite genome, and the evolutionary forces that have led to their high frequency and strong linkage disequilibrium in African P. falciparum populations.

We conducted a genome-wide association study of host resistance to severePlasmodium falciparummalaria in over 17,000 individuals from 11 malaria-endemic countries, undertaking a wide ranging analysis which identifies five replicable associations with genome-wide levels of evidence. Our findings include a newly implicated variant on chromosome 6 associated with risk of cerebral malaria, and the discovery of an erythroid-specific transcription start site underlying the association inATP2B4. Previously reported HLA associations cannot be replicated in this dataset. We estimate substantial heritability of severe malaria (h2~ 23%), of which around 10% is explained by the currently identified associations. Our dataset will provide a major building block for future research on the genetic determinants of disease in these diverse human populations.

INTRODUCTION Malaria parasites cause human disease by invading and replicating inside red blood cells. In the case of Plasmodium falciparum , this can lead to severe forms of malaria that are a major cause of childhood mortality in Africa. This species of parasite enters the red blood cell through interactions with surface proteins including the glycophorins GYPA and GYPB, which determine the polymorphic MNS blood group system. In a recent genome-wide association study, we identified alleles associated with protection against severe malaria near the cluster of genes encoding these invasion receptors. RATIONALE Investigation of genetic variants at this locus and their relation to severe malaria is challenging because of the high sequence similarity between the neighboring glycophorin genes and the relative lack of available sequence data capturing the genetic diversity of sub-Saharan Africa. To better assess whether variation in the glycophorin genes could explain the signal of association, we generated additional sequence data from sub-Saharan African populations and developed an analytical approach to characterize structural variation at this complex locus. RESULTS Using 765 newly sequenced human genomes from 10 African ethnic groups along with data from the 1000 Genomes Project, we generated a reference panel of haplotypes across the glycophorin region. In addition to single-nucleotide polymorphisms and short indels, we assayed large copy number variants (CNVs) using sequencing read depth and uncovered extensive structural diversity. By imputing from this reference panel into 4579 severe malaria cases and 5310 controls from three African populations, we found that a complex CNV, here called DUP4, is associated with resistance to severe malaria and fully explains the previously reported signal of association. In our sample, DUP4 is present only in east Africa, and this localization, as well as the extent of similarity between DUP4 haplotypes, suggests that it has recently increased in frequency, presumably under natural selection due to malaria. To evaluate the potential functional consequences of this structural variant, we analyzed high-coverage sequence-read data from multiple individuals to generate a model of the DUP4 chromosome structure. The DUP4 haplotype contains five glycophorin genes, including two hybrid genes that juxtapose the extracellular domain of GYPB with the transmembrane and intracellular domains of GYPA. Noting that these predicted hybrids are characteristic of the Dantu antigen in the MNS blood group system, we sequenced a Dantu positive individual and confirmed that DUP4 is the molecular basis of the Dantu NE blood group variant. CONCLUSION Although a role for GYPA and GYPB in parasite invasion is well known, a direct link between glycophorin polymorphisms and clinical susceptibility to malaria has been elusive. Here we have provided a systematic catalog of CNVs, describing structural diversity that may have functional importance at this locus. Our results identify a specific variant that encodes hybrid glycophorin proteins and is associated with protection against severe malaria. This discovery calls for further work to determine how this particular molecular rearrangement affects parasite invasion and the red blood cell response and may lead us toward new parasite vulnerabilities that can be utilized in future interventions against this deadly disease.

The wide variety of disease associations reported at the TNF locus raises the question of how much variation exists within a single population. To address this question, we sequenced the entire TNF gene in 72 chromosomes from healthy residents of a village in The Gambia, West Africa. We found 12 polymorphisms in 4393 nucleotides, of which five have not been previously described, giving an estimated nucleotide diversity (theta) of 5.6 x 10(-4). A significantly higher frequency of polymorphisms was found in the promoter region than in the coding region (8/1256 vs 0/882 nucleotides, P = 0.02). All polymorphisms with the exception of one rare allele were found to be present in Malawi, which is both geographically and genetically distant from The Gambia. Genotyping of 424 Gambian and 121 Malawian adults showed a significant frequency difference between the two populations for eight of the 12 polymorphisms, but the average fixation index across the variable sites was relatively low (F(ST) = 0.007). We conclude that, at the TNF locus, the nucleotide diversity found within a single African village is similar to the global value for human autosomal genes sampled across different continents.

Interferon-gamma, encoded by IFNG, is a key immunological mediator that is believed to play both a protective and a pathological role in malaria. Here, we investigate the relationship between IFNG variation and susceptibility to malaria. We began by analysing West African and European haplotype structure and patterns of linkage disequilibrium across a 100 kb genomic region encompassing IFNG and its immediate neighbours IL22 and IL26. A large case-control study of severe malaria in a West Africa population identified several weak associations with individual single-nucleotide polymorphisms in the IFNG and IL22 genes, and defined two IL22 haplotypes that are, respectively, associated with resistance and susceptibility. These data provide a starting point for functional and genetic analysis of the IFNG genomic region in malaria and other infectious and inflammatory conditions affecting African populations.

The 1000 Genomes Project set out to provide a comprehensive description of common human genetic variation by applying whole-genome sequencing to a diverse set of individuals from multiple populations. Here we report completion of the project, having reconstructed the genomes of 2,504 individuals from 26 populations using a combination of low-coverage whole-genome sequencing, deep exome sequencing, and dense microarray genotyping. We characterized a broad spectrum of genetic variation, in total over 88 million variants (84.7 million single nucleotide polymorphisms (SNPs), 3.6 million short insertions/deletions (indels), and 60,000 structural variants), all phased onto high-quality haplotypes. This resource includes >99% of SNP variants with a frequency of >1% for a variety of ancestries. We describe the distribution of genetic variation across the global sample, and discuss the implications for common disease studies., Wellcome Trust (London, England) (Core Award 090532/Z/09/Z), Wellcome Trust (London, England) (Senior Investigator Award 095552/Z/11/Z ), Wellcome Trust (London, England) (WT095908), Wellcome Trust (London, England) (WT109497), Wellcome Trust (London, England) (WT098051), Wellcome Trust (London, England) (WT086084/Z/08/Z), Wellcome Trust (London, England) (WT100956/Z/13/Z ), Wellcome Trust (London, England) (WT097307), Wellcome Trust (London, England) (WT0855322/Z/08/Z ), Wellcome Trust (London, England) (WT090770/Z/09/Z ), Wellcome Trust (London, England) (Major Overseas program in Vietnam grant 089276/Z.09/Z), Medical Research Council (Great Britain) (grant G0801823), Biotechnology and Biological Sciences Research Council (Great Britain) (grant BB/I02593X/1), Biotechnology and Biological Sciences Research Council (Great Britain) (grant BB/I021213/1), Zhongguo ke xue ji shu qing bao yan jiu suo. Office of 863 Programme of China (2012AA02A201), National Basic Research Program of China (2011CB809201), National Basic Research Program of China (2011CB809202), National Basic Research Program of China (2011CB809203), National Natural Science Foundation of China (31161130357), Shenzhen Municipal Government of China (grant ZYC201105170397A), Canadian Institutes of Health Research (grant 136855), Quebec Ministry of Economic Development, Innovation, and Exports (PSR-SIIRI-195), Germany. Bundesministerium für Bildung und Forschung (0315428A), Germany. Bundesministerium für Bildung und Forschung (01GS08201), Germany. Bundesministerium für Bildung und Forschung (BMBF-EPITREAT grant 0316190A), Deutsche Forschungsgemeinschaft (Emmy Noether Grant KO4037/1-1), Beatriu de Pinos Program (2006 BP-A 10144), Beatriu de Pinos Program (2009 BP-B 00274), Spanish National Institute for Health (grant PRB2 IPT13/0001-ISCIII-SGEFI/FEDER), Japan Society for the Promotion of Science (fellowship number PE13075), Marie Curie Actions Career Integration (grant 303772), Fonds National Suisse del la Recherche, SNSF, Scientifique (31003A_130342), National Center for Biotechnology Information (U.S.) (U54HG3067), National Center for Biotechnology Information (U.S.) (U54HG3273), National Center for Biotechnology Information (U.S.) (U01HG5211), National Center for Biotechnology Information (U.S.) (U54HG3079), National Center for Biotechnology Information (U.S.) (R01HG2898), National Center for Biotechnology Information (U.S.) (R01HG2385), National Center for Biotechnology Information (U.S.) (RC2HG5552), National Center for Biotechnology Information (U.S.) (U01HG6513), National Center for Biotechnology Information (U.S.) (U01HG5214), National Center for Biotechnology Information (U.S.) (U01HG5715), National Center for Biotechnology Information (U.S.) (U01HG5718), National Center for Biotechnology Information (U.S.) (U01HG5728), National Center for Biotechnology Information (U.S.) (U41HG7635), National Center for Biotechnology Information (U.S.) (U41HG7497), National Center for Biotechnology Information (U.S.) (R01HG4960), National Center for Biotechnology Information (U.S.) (R01HG5701), National Center for Biotechnology Information (U.S.) (R01HG5214), National Center for Biotechnology Information (U.S.) (R01HG6855), National Center for Biotechnology Information (U.S.) (R01HG7068), National Center for Biotechnology Information (U.S.) (R01HG7644), National Center for Biotechnology Information (U.S.) (DP2OD6514), National Center for Biotechnology Information (U.S.) (DP5OD9154), National Center for Biotechnology Information (U.S.) (R01CA166661), National Center for Biotechnology Information (U.S.) (R01CA172652), National Center for Biotechnology Information (U.S.) (P01GM99568), National Center for Biotechnology Information (U.S.) (R01GM59290), National Center for Biotechnology Information (U.S.) (R01GM104390), National Center for Biotechnology Information (U.S.) (T32GM7790), National Center for Biotechnology Information (U.S.) (R01HL87699), National Center for Biotechnology Information (U.S.) (R01HL104608), National Center for Biotechnology Information (U.S.) (T32HL94284), National Center for Biotechnology Information (U.S.) (HHSN268201100040C), National Center for Biotechnology Information (U.S.) (HHSN272201000025C), Lundbeck Foundation (grant R170-2014-1039, Simons Foundation (SFARI award SF51), National Science Foundation (U.S.) (Research Fellowship DGE-1147470)