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    An open dataset of Plasmodium vivax genome variation in 1,895 worldwide samples.
    MalariaGEN, ; Adam, I ; Alam, MS ; Alemu, S ; Amaratunga, C ; Amato, R ; Andrianaranjaka, V ; Anstey, NM ; Aseffa, A ; Ashley, E ; Assefa, A ; Auburn, S ; Barber, BE ; Barry, A ; Batista Pereira, D ; Cao, J ; Chau, NH ; Chotivanich, K ; Chu, C ; Dondorp, AM ; Drury, E ; Echeverry, DF ; Erko, B ; Espino, F ; Fairhurst, R ; Faiz, A ; Fernanda Villegas, M ; Gao, Q ; Golassa, L ; Goncalves, S ; Grigg, MJ ; Hamedi, Y ; Hien, TT ; Htut, Y ; Johnson, KJ ; Karunaweera, N ; Khan, W ; Krudsood, S ; Kwiatkowski, DP ; Lacerda, M ; Ley, B ; Lim, P ; Liu, Y ; Llanos-Cuentas, A ; Lon, C ; Lopera-Mesa, T ; Marfurt, J ; Michon, P ; Miotto, O ; Mohammed, R ; Mueller, I ; Namaik-Larp, C ; Newton, PN ; Nguyen, T-N ; Nosten, F ; Noviyanti, R ; Pava, Z ; Pearson, RD ; Petros, B ; Phyo, AP ; Price, RN ; Pukrittayakamee, S ; Rahim, AG ; Randrianarivelojosia, M ; Rayner, JC ; Rumaseb, A ; Siegel, SV ; Simpson, VJ ; Thriemer, K ; Tobon-Castano, A ; Trimarsanto, H ; Urbano Ferreira, M ; Vélez, ID ; Wangchuk, S ; Wellems, TE ; White, NJ ; William, T ; Yasnot, MF ; Yilma, D (F1000 Research Ltd, 2022)
    This report describes the MalariaGEN Pv4 dataset, a new release of curated genome variation data on 1,895 samples of Plasmodium vivax collected at 88 worldwide locations between 2001 and 2017. It includes 1,370 new samples contributed by MalariaGEN and VivaxGEN partner studies in addition to previously published samples from these and other sources. We provide genotype calls at over 4.5 million variable positions including over 3 million single nucleotide polymorphisms (SNPs), as well as short indels and tandem duplications. This enlarged dataset highlights major compartments of parasite population structure, with clear differentiation between Africa, Latin America, Oceania, Western Asia and different parts of Southeast Asia. Each sample has been classified for drug resistance to sulfadoxine, pyrimethamine and mefloquine based on known markers at the dhfr, dhps and mdr1 loci. The prevalence of all of these resistance markers was much higher in Southeast Asia and Oceania than elsewhere. This open resource of analysis-ready genome variation data from the MalariaGEN and VivaxGEN networks is driven by our collective goal to advance research into the complex biology of P. vivax and to accelerate genomic surveillance for malaria control and elimination.
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    An open dataset of Plasmodium falciparum genome variation in 7,000 worldwide samples.
    MalariaGEN, ; Ahouidi, A ; Ali, M ; Almagro-Garcia, J ; Amambua-Ngwa, A ; Amaratunga, C ; Amato, R ; Amenga-Etego, L ; Andagalu, B ; Anderson, TJC ; Andrianaranjaka, V ; Apinjoh, T ; Ariani, C ; Ashley, EA ; Auburn, S ; Awandare, GA ; Ba, H ; Baraka, V ; Barry, AE ; Bejon, P ; Bertin, GI ; Boni, MF ; Borrmann, S ; Bousema, T ; Branch, O ; Bull, PC ; Busby, GBJ ; Chookajorn, T ; Chotivanich, K ; Claessens, A ; Conway, D ; Craig, A ; D'Alessandro, U ; Dama, S ; Day, NP ; Denis, B ; Diakite, M ; Djimdé, A ; Dolecek, C ; Dondorp, AM ; Drakeley, C ; Drury, E ; Duffy, P ; Echeverry, DF ; Egwang, TG ; Erko, B ; Fairhurst, RM ; Faiz, A ; Fanello, CA ; Fukuda, MM ; Gamboa, D ; Ghansah, A ; Golassa, L ; Goncalves, S ; Hamilton, WL ; Harrison, GLA ; Hart, L ; Henrichs, C ; Hien, TT ; Hill, CA ; Hodgson, A ; Hubbart, C ; Imwong, M ; Ishengoma, DS ; Jackson, SA ; Jacob, CG ; Jeffery, B ; Jeffreys, AE ; Johnson, KJ ; Jyothi, D ; Kamaliddin, C ; Kamau, E ; Kekre, M ; Kluczynski, K ; Kochakarn, T ; Konaté, A ; Kwiatkowski, DP ; Kyaw, MP ; Lim, P ; Lon, C ; Loua, KM ; Maïga-Ascofaré, O ; Malangone, C ; Manske, M ; Marfurt, J ; Marsh, K ; Mayxay, M ; Miles, A ; Miotto, O ; Mobegi, V ; Mokuolu, OA ; Montgomery, J ; Mueller, I ; Newton, PN ; Nguyen, T ; Nguyen, T-N ; Noedl, H ; Nosten, F ; Noviyanti, R ; Nzila, A ; Ochola-Oyier, LI ; Ocholla, H ; Oduro, A ; Omedo, I ; Onyamboko, MA ; Ouedraogo, J-B ; Oyebola, K ; Pearson, RD ; Peshu, N ; Phyo, AP ; Plowe, CV ; Price, RN ; Pukrittayakamee, S ; Randrianarivelojosia, M ; Rayner, JC ; Ringwald, P ; Rockett, KA ; Rowlands, K ; Ruiz, L ; Saunders, D ; Shayo, A ; Siba, P ; Simpson, VJ ; Stalker, J ; Su, X-Z ; Sutherland, C ; Takala-Harrison, S ; Tavul, L ; Thathy, V ; Tshefu, A ; Verra, F ; Vinetz, J ; Wellems, TE ; Wendler, J ; White, NJ ; Wright, I ; Yavo, W ; Ye, H (F1000 Research Ltd, 2021)
    MalariaGEN is a data-sharing network that enables groups around the world to work together on the genomic epidemiology of malaria. Here we describe a new release of curated genome variation data on 7,000 Plasmodium falciparum samples from MalariaGEN partner studies in 28 malaria-endemic countries. High-quality genotype calls on 3 million single nucleotide polymorphisms (SNPs) and short indels were produced using a standardised analysis pipeline. Copy number variants associated with drug resistance and structural variants that cause failure of rapid diagnostic tests were also analysed.  Almost all samples showed genetic evidence of resistance to at least one antimalarial drug, and some samples from Southeast Asia carried markers of resistance to six commonly-used drugs. Genes expressed during the mosquito stage of the parasite life-cycle are prominent among loci that show strong geographic differentiation. By continuing to enlarge this open data resource we aim to facilitate research into the evolutionary processes affecting malaria control and to accelerate development of the surveillance toolkit required for malaria elimination.
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    Global diversity and balancing selection of 23 leading Plasmodium falciparum candidate vaccine antigens.
    Naung, MT ; Martin, E ; Munro, J ; Mehra, S ; Guy, AJ ; Laman, M ; Harrison, GLA ; Tavul, L ; Hetzel, M ; Kwiatkowski, D ; Mueller, I ; Bahlo, M ; Barry, AE ; Wallqvist, A (Public Library of Science (PLoS), 2022-02)
    Investigation of the diversity of malaria parasite antigens can help prioritize and validate them as vaccine candidates and identify the most common variants for inclusion in vaccine formulations. Studies of vaccine candidates of the most virulent human malaria parasite, Plasmodium falciparum, have focused on a handful of well-known antigens, while several others have never been studied. Here we examine the global diversity and population structure of leading vaccine candidate antigens of P. falciparum using the MalariaGEN Pf3K (version 5.1) resource, comprising more than 2600 genomes from 15 malaria endemic countries. A stringent variant calling pipeline was used to extract high quality antigen gene 'haplotypes' from the global dataset and a new R-package named VaxPack was used to streamline population genetic analyses. In addition, a newly developed algorithm that enables spatial averaging of selection pressure on 3D protein structures was applied to the dataset. We analysed the genes encoding 23 leading and novel candidate malaria vaccine antigens including csp, trap, eba175, ama1, rh5, and CelTOS. Our analysis shows that current malaria vaccine formulations are based on rare haplotypes and thus may have limited efficacy against natural parasite populations. High levels of diversity with evidence of balancing selection was detected for most of the erythrocytic and pre-erythrocytic antigens. Measures of natural selection were then mapped to 3D protein structures to predict targets of functional antibodies. For some antigens, geographical variation in the intensity and distribution of these signals on the 3D structure suggests adaptation to different human host or mosquito vector populations. This study provides an essential framework for the diversity of P. falciparum antigens to be considered in the design of the next generation of malaria vaccines.
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    Surveillance of molecular markers of Plasmodium falciparum artemisinin resistance (kelch13 mutations) in Papua New Guinea between 2016 and 2018
    Lautu-Gumal, D ; Razook, Z ; Koleala, T ; Nate, E ; McEwen, S ; Timbi, D ; Hetzel, MW ; Lavu, E ; Tefuarani, N ; Makita, L ; Kazura, J ; Mueller, I ; Pomat, W ; Laman, M ; Robinson, LJ ; Barry, AE (ELSEVIER SCI LTD, 2021-07-13)
    Plasmodium falciparum resistance to artemisinin-based combination therapy (ACT) is a global threat to malaria control and elimination efforts. Mutations in the P. falciparum kelch13 gene (Pfk13) that are associated with delayed parasite clearance have emerged on the Thai-Cambodian border since 2008. There is growing evidence of widespread Pfk13 mutations throughout South-East Asia and they have independently emerged in other endemic regions. In Papua New Guinea (PNG), Pfk13 "C580Y" mutant parasites with reduced in vitro sensitivity to artemisinin have been isolated in Wewak, a port town in East Sepik Province. However, the extent of any local spread of these mutant parasites in other parts of PNG is unknown. We investigated the prevalence of Pfk13 mutations in multiple malaria-endemic regions of PNG. P. falciparum isolates (n = 1152) collected between 2016 and 2018 and assessed for Pfk13 variation by sequencing. Of 663 high quality Pfk13 sequences a total of five variants were identified. They included C580Y, a mutation at a previously documented polymorphic locus: N499K, and three previously undescribed mutations: R471C, K586E and Y635C. All variants were found in single isolates, indicating that these Pfk13 mutations were rare in the areas surveyed. Notably, C580Y was absent from Maprik district, which neighbours Wewak where C580Y mutant parasites were previously identified. The single C580Y isolate was found in the port town of Lae, Morobe Province, a potential entry site for the importation of drug resistant parasites into PNG. Although sample size in this location was small (n = 5), our identification of a C580Y mutant in this second location is concerning, highlighting the urgent need for further surveillance in Lae. Other Pfk13 mutants were rare in PNG between 2016 and 2018. Continued surveillance for molecular markers of drug resistance is critically important to inform malaria control in PNG.
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    MonitoringPlasmodium falciparumandPlasmodium vivaxusing microsatellite markers indicates limited changes in population structure after substantial transmission decline in Papua New Guinea
    Kattenberg, JH ; Razook, Z ; Keo, R ; Koepfli, C ; Jennison, C ; Lautu-Gumal, D ; Fola, AA ; Ome-Kaius, M ; Barnadas, C ; Siba, P ; Felger, I ; Kazura, J ; Mueller, I ; Robinson, LJ ; Barry, AE (WILEY, 2020-10-16)
    Monitoring the genetic structure of pathogen populations may be an economical and sensitive approach to quantify the impact of control on transmission dynamics, highlighting the need for a better understanding of changes in population genetic parameters as transmission declines. Here we describe the first population genetic analysis of two major human malaria parasites, Plasmodium falciparum (Pf) and Plasmodium vivax (Pv), following nationwide distribution of long-lasting insecticide-treated nets (LLINs) in Papua New Guinea (PNG). Parasite isolates from pre- (2005-2006) and post-LLIN (2010-2014) were genotyped using microsatellite markers. Despite parasite prevalence declining substantially (East Sepik Province: Pf = 54.9%-8.5%, Pv = 35.7%-5.6%, Madang Province: Pf = 38.0%-9.0%, Pv: 31.8%-19.7%), genetically diverse and intermixing parasite populations remained. Pf diversity declined modestly post-LLIN relative to pre-LLIN (East Sepik: Rs  = 7.1-6.4, HE  = 0.77-0.71; Madang: Rs  = 8.2-6.1, HE  = 0.79-0.71). Unexpectedly, population structure present in pre-LLIN populations was lost post-LLIN, suggesting that more frequent human movement between provinces may have contributed to higher gene flow. Pv prevalence initially declined but increased again in one province, yet diversity remained high throughout the study period (East Sepik: Rs  = 11.4-9.3, HE  = 0.83-0.80; Madang: Rs  = 12.2-14.5, HE  = 0.85-0.88). Although genetic differentiation values increased between provinces over time, no significant population structure was observed at any time point. For both species, a decline in multiple infections and increasing clonal transmission and significant multilocus linkage disequilibrium post-LLIN were positive indicators of impact on the parasite population using microsatellite markers. These parameters may be useful adjuncts to traditional epidemiological tools in the early stages of transmission reduction.
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    Emergence of artemisinin-resistant Plasmodium falciparum with kelch13 C580Y mutations on the island of New Guinea
    Miotto, O ; Sekihara, M ; Tachibana, S-I ; Yamauchi, M ; Pearson, RD ; Amato, R ; Goncalves, S ; Mehra, S ; Noviyanti, R ; Marfurt, J ; Auburn, S ; Price, RN ; Mueller, I ; Ikeda, M ; Mori, T ; Hirai, M ; Tavul, L ; Hetzel, MW ; Laman, M ; Barry, AE ; Ringwald, P ; Ohashi, J ; Hombhanje, F ; Kwiatkowski, DP ; Mita, T ; Billker, O (PUBLIC LIBRARY SCIENCE, 2020-12-01)
    The rapid and aggressive spread of artemisinin-resistant Plasmodium falciparum carrying the C580Y mutation in the kelch13 gene is a growing threat to malaria elimination in Southeast Asia, but there is no evidence of their spread to other regions. We conducted cross-sectional surveys in 2016 and 2017 at two clinics in Wewak, Papua New Guinea (PNG) where we identified three infections caused by C580Y mutants among 239 genotyped clinical samples. One of these mutants exhibited the highest survival rate (6.8%) among all parasites surveyed in ring-stage survival assays (RSA) for artemisinin. Analyses of kelch13 flanking regions, and comparisons of deep sequencing data from 389 clinical samples from PNG, Indonesian Papua and Western Cambodia, suggested an independent origin of the Wewak C580Y mutation, showing that the mutants possess several distinctive genetic features. Identity by descent (IBD) showed that multiple portions of the mutants' genomes share a common origin with parasites found in Indonesian Papua, comprising several mutations within genes previously associated with drug resistance, such as mdr1, ferredoxin, atg18 and pnp. These findings suggest that a P. falciparum lineage circulating on the island of New Guinea has gradually acquired a complex ensemble of variants, including kelch13 C580Y, which have affected the parasites' drug sensitivity. This worrying development reinforces the need for increased surveillance of the evolving parasite populations on the island, to contain the spread of resistance.
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    Molecular epidemiology of residual Plasmodium vivax transmission in a paediatric cohort in Solomon Islands
    Quah, YW ; Waltmann, A ; Karl, S ; White, MT ; Vahi, V ; Darcy, A ; Pitakaka, F ; Whittaker, M ; Tisch, DJ ; Barry, A ; Barnadas, C ; Kazura, J ; Mueller, I (BMC, 2019-03-28)
    BACKGROUND: Following the scale-up of intervention efforts, malaria burden has decreased dramatically in Solomon Islands (SI). Submicroscopic and asymptomatic Plasmodium vivax infections are now the major challenge for malaria elimination in this country. Since children have higher risk of contracting malaria, this study investigated the dynamics of Plasmodium spp. infections among children including the associated risk factors of residual P. vivax burden. METHODS: An observational cohort study was conducted among 860 children aged 0.5-12 years in Ngella (Central Islands Province, SI). Children were monitored by active and passive surveillances for Plasmodium spp. infections and illness. Parasites were detected by quantitative real-time PCR (qPCR) and genotyped. Comprehensive statistical analyses of P. vivax infection prevalence, molecular force of blood stage infection (molFOB) and infection density were conducted. RESULTS: Plasmodium vivax infections were common (overall prevalence: 11.9%), whereas Plasmodium falciparum infections were rare (0.3%) but persistent. Although children acquire an average of 1.1 genetically distinct P. vivax blood-stage infections per year, there was significant geographic heterogeneity in the risks of P. vivax infections across Ngella (prevalence: 1.2-47.4%, p < 0.01; molFOB: 0.05-4.6/year, p < 0.01). Malaria incidence was low (IR: 0.05 episodes/year-at-risk). Age and measures of high exposure were the key risk factors for P. vivax infections and disease. Malaria incidence and infection density decreased with age, indicating significant acquisition of immunity. G6PD deficient children (10.8%) that did not receive primaquine treatment had a significantly higher prevalence (aOR: 1.77, p = 0.01) and increased risk of acquiring new bloodstage infections (molFOB aIRR: 1.51, p = 0.03), underscoring the importance of anti-relapse treatment. CONCLUSION: Residual malaria transmission in Ngella exhibits strong heterogeneity and is characterized by a high proportion of submicroscopic and asymptomatic P. vivax infections, alongside sporadic P. falciparum infections. Implementing an appropriate primaquine treatment policy to prevent P. vivax relapses and specific targeting of control interventions to high risk areas will be required to accelerate ongoing control and elimination activities.
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    Sustained Malaria Control Over an 8-Year Period in Papua New Guinea: The Challenge of Low-Density Asymptomatic Plasmodium Infections
    Koepfli, C ; Ome-Kaius, M ; Jally, S ; Malau, E ; Maripal, S ; Ginny, J ; Timinao, L ; Kattenberg, JH ; Obadia, T ; White, M ; Rarau, P ; Senn, N ; Barry, AE ; Kazura, JW ; Mueller, I ; Robinson, LJ (OXFORD UNIV PRESS INC, 2017-12-01)
    Background: The scale-up of effective malaria control in the last decade has resulted in a substantial decline in the incidence of clinical malaria in many countries. The effects on the proportions of asymptomatic and submicroscopic infections and on transmission potential are yet poorly understood. Methods: In Papua New Guinea, vector control has been intensified since 2008, and improved diagnosis and treatment was introduced in 2012. Cross-sectional surveys were conducted in Madang Province in 2006 (with 1280 survey participants), 2010 (with 2117 participants), and 2014 (with 2516 participants). Infections were quantified by highly sensitive quantitative polymerase chain reaction (PCR) analysis, and gametocytes were quantified by reverse-transcription qPCR analysis. Results: Plasmodium falciparum prevalence determined by qPCR decreased from 42% in 2006 to 9% in 2014. The P. vivax prevalence decreased from 42% in 2006 to 13% in 2010 but then increased to 20% in 2014. Parasite densities decreased 5-fold from 2006 to 2010; 72% of P. falciparum and 87% of P. vivax infections were submicroscopic in 2014. Gametocyte density and positivity correlated closely with parasitemia, and population gametocyte prevalence decreased 3-fold for P. falciparum and 29% for P. vivax from 2010 to 2014. Conclusions: Sustained control has resulted in reduced malaria transmission potential, but an increasing proportion of gametocyte carriers are asymptomatic and submicroscopic and represent a challenge to malaria control.
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    Genomic analysis of local variation and recent evolution in Plasmodium vivax
    Pearson, RD ; Amato, R ; Auburn, S ; Miotto, O ; Almagro-Garcia, J ; Amaratunga, C ; Suon, S ; Mao, S ; Noviyanti, R ; Trimarsanto, H ; Marfurt, J ; Anstey, NM ; William, T ; Boni, MF ; Dolecek, C ; Hien, TT ; White, NJ ; Michon, P ; Siba, P ; Tavul, L ; Harrison, G ; Barry, A ; Mueller, I ; Ferreira, MU ; Karunaweera, N ; Randrianarivelojosia, M ; Gao, Q ; Hubbart, C ; Hart, L ; Jeffery, B ; Drury, E ; Mead, D ; Kekre, M ; Campino, S ; Manske, M ; Cornelius, VJ ; MacInnis, B ; Rockett, KA ; Miles, A ; Rayner, JC ; Fairhurst, RM ; Nosten, F ; Price, RN ; Kwiatkowski, DP (NATURE PUBLISHING GROUP, 2016-08-01)
    The widespread distribution and relapsing nature of Plasmodium vivax infection present major challenges for the elimination of malaria. To characterize the genetic diversity of this parasite in individual infections and across the population, we performed deep genome sequencing of >200 clinical samples collected across the Asia-Pacific region and analyzed data on >300,000 SNPs and nine regions of the genome with large copy number variations. Individual infections showed complex patterns of genetic structure, with variation not only in the number of dominant clones but also in their level of relatedness and inbreeding. At the population level, we observed strong signals of recent evolutionary selection both in known drug resistance genes and at new loci, and these varied markedly between geographical locations. These findings demonstrate a dynamic landscape of local evolutionary adaptation in the parasite population and provide a foundation for genomic surveillance to guide effective strategies for control and elimination of P. vivax.
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    Genomic epidemiology of artemisinin resistant malaria
    Amato, R ; Miotto, O ; Woodrow, CJ ; Almagro-Garcia, J ; Sinha, I ; Campino, S ; Mead, D ; Drury, E ; Kekre, M ; Sanders, M ; Amambua-Ngwa, A ; Amaratunga, C ; Amenga-Etego, L ; Andrianaranjaka, V ; Apinjoh, T ; Ashley, E ; Auburn, S ; Awandare, GA ; Baraka, V ; Barry, A ; Boni, MF ; Borrmann, S ; Bousema, T ; Branch, O ; Bull, PC ; Chotivanich, K ; Conway, DJ ; Craig, A ; Day, NP ; Djimde, A ; Dolecek, C ; Dondorp, AM ; Drakeley, C ; Duffy, P ; Echeverry, DF ; Egwang, TG ; Fairhurst, RM ; Faiz, MA ; Fanello, CI ; Tran, TH ; Hodgson, A ; Imwong, M ; Ishengoma, D ; Lim, P ; Lon, C ; Marfurt, J ; Marsh, K ; Mayxay, M ; Michon, P ; Mobegi, V ; Mokuolu, OA ; Montgomery, J ; Mueller, I ; Kyaw, MP ; Newton, PN ; Nosten, F ; Noviyanti, R ; Nzila, A ; Ocholla, H ; Oduro, A ; Onyamboko, M ; Ouedraogo, J-B ; Phyo, APP ; Plowe, C ; Price, RN ; Pukrittayakamee, S ; Randrianarivelojosia, M ; Ringwald, P ; Ruiz, L ; Saunders, D ; Shayo, A ; Siba, P ; Takala-Harrison, S ; Thanh, T-NN ; Thathy, V ; Verra, F ; Wendler, J ; White, NJ ; Ye, H ; Cornelius, VJ ; Giacomantonio, R ; Muddyman, D ; Henrichs, C ; Malangone, C ; Jyothi, D ; Pearson, RD ; Rayner, JC ; McVean, G ; Rockett, KA ; Miles, A ; Vauterin, P ; Jeffery, B ; Manske, M ; Stalker, J ; Maclnnis, B ; Kwiatkowski, DP (ELIFE SCIENCES PUBLICATIONS LTD, 2016-03-04)
    The current epidemic of artemisinin resistant Plasmodium falciparum in Southeast Asia is the result of a soft selective sweep involving at least 20 independent kelch13 mutations. In a large global survey, we find that kelch13 mutations which cause resistance in Southeast Asia are present at low frequency in Africa. We show that African kelch13 mutations have originated locally, and that kelch13 shows a normal variation pattern relative to other genes in Africa, whereas in Southeast Asia there is a great excess of non-synonymous mutations, many of which cause radical amino-acid changes. Thus, kelch13 is not currently undergoing strong selection in Africa, despite a deep reservoir of variations that could potentially allow resistance to emerge rapidly. The practical implications are that public health surveillance for artemisinin resistance should not rely on kelch13 data alone, and interventions to prevent resistance must account for local evolutionary conditions, shown by genomic epidemiology to differ greatly between geographical regions.