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Malaria parasites break down host haemoglobin into peptides and amino acids in the digestive vacuole for export to the parasite cytoplasm for growth: interrupting this process is central to the mode of action of several antimalarial drugs. Mutations in the chloroquine (CQ) resistance transporter, pfcrt, located in the digestive vacuole membrane, confer CQ resistance in Plasmodium falciparum, and typically also affect parasite fitness. However, the role of other parasite loci in the evolution of CQ resistance is unclear. Here we use a combination of population genomics, genetic crosses and gene editing to demonstrate that a second vacuolar transporter plays a key role in both resistance and compensatory evolution. Longitudinal genomic analyses of the Gambian parasites revealed temporal signatures of selection on a putative amino acid transporter (pfaat1) variant S258L, which increased from 0% to 97% in frequency between 1984 and 2014 in parallel with the pfcrt1 K76T variant. Parasite genetic crosses then identified a chromosome 6 quantitative trait locus containing pfaat1 that is selected by CQ treatment. Gene editing demonstrated that pfaat1 S258L potentiates CQ resistance but at a cost of reduced fitness, while pfaat1 F313S, a common southeast Asian polymorphism, reduces CQ resistance while restoring fitness. Our analyses reveal hidden complexity in CQ resistance evolution, suggesting that pfaat1 may underlie regional differences in the dynamics of resistance evolution, and modulate parasite resistance or fitness by manipulating the balance between both amino acid and drug transport.

With global progress towards malaria reduction stalling, further analysis of the epidemiology is required, particularly in countries with the highest burden. National surveys have mostly analysed infection prevalence, while large-scale data on parasite density and different developmental forms are rarely available. In Nigeria, the country with the largest burden globally, blood slide microscopy of children up to five years of age was conducted in the 2018 National Demographic and Health Survey, and infection prevalence previously reported. In the current study, malaria parasite density measurements are reported and analysed for 7783 of the children sampled across the 36 states within the six geopolitical zones of the country, with asexual and sexual stages considered separately and comparisons of infections with different malaria parasite species being performed. Across all states of Nigeria, there was a positive correlation between mean asexual parasite density in peripheral blood of infected individuals and prevalence of infection in the community (Spearman’s rho = 0.39, P = 0.02). Asexual parasite densities were highest in the northern geopolitical zones (geometric means > 2000 μL-1), extending the evidence of exceptionally high infection burden in many areas. Sexual parasite prevalence in each state was highly correlated with asexual parasite prevalence (Spearman’s rho = 0.70, P < 0.001), although sexual parasite densities were low (geometric means < 100 μL-1in all zones). Infants had lower parasite densities than children above one year of age, but there were no differences between male and female children. Most infections were ofP. falciparum, which had higher asexual densities but lower sexual parasite densities thanP. malariaeorP. ovalemono-infections. However, mixed species infections had the highest asexual parasite densities. It is recommended that future large surveys in high burden countries measure parasite densities as well as developmental stages and species, to improve the quality of malaria epidemiology and tracking of future changes.

BACKGROUND: Malaria control interventions have led to a decline in transmission intensity in many endemic areas, and resulted in elimination in some areas. This decline, however, will lead to delayed acquisition of protective immunity and thus impact disease manifestation and outcomes. Therefore, the variation in clinical and haematological parameters in children with malaria was assessed across three areas in Ghana with varying transmission intensities. METHODS: A total of 568 children between the ages of 2 and 14 years with confirmed malaria were recruited in hospitals in three areas with varying transmission intensities (Kintampo > Navrongo > Accra) and a comprehensive analysis of parasitological, clinical, haematological and socio-economic parameters was performed. RESULTS: Areas of lower malaria transmission tended to have lower disease severity in children with malaria, characterized by lower parasitaemias and higher haemoglobin levels. In addition, total white cell counts and percent lymphocytes decreased with decreasing transmission intensity. The heterozygous sickle haemoglobin genotype was protective against disease severity in Kintampo (P = 0.016), although this was not significant in Accra and Navrongo. Parasitaemia levels were not a significant predictor of haemoglobin level after controlling for age and gender. However, higher haemoglobin levels in children were associated with certain socioeconomic factors, such as having fathers who had any type of employment (P

Many human genetic associations with resistance to malaria have been reported, but few have been reliably replicated. We collected data on 11,890 cases of severe malaria due to Plasmodium falciparum and 17,441 controls from 12 locations in Africa, Asia and Oceania. We tested 55 SNPs in 27 loci previously reported to associate with severe malaria. There was evidence of association at P < 1 × 10(-4) with the HBB, ABO, ATP2B4, G6PD and CD40LG loci, but previously reported associations at 22 other loci did not replicate in the multicenter analysis. The large sample size made it possible to identify authentic genetic effects that are heterogeneous across populations or phenotypes, with a striking example being the main African form of G6PD deficiency, which reduced the risk of cerebral malaria but increased the risk of severe malarial anemia. The finding that G6PD deficiency has opposing effects on different fatal complications of P. falciparum infection indicates that the evolutionary origins of this common human genetic disorder are more complex than previously supposed.

Background Genome wide sequence analyses of malaria parasites from widely separated areas of the world have identified contrasting population structures and signatures of selection. To compare relatively closely situated but ecologically contrasting regions within an endemic African country, population samples of Plasmodium falciparum clinical isolates were collected in Ghana from Kintampo in the central forest-savannah area, and Navrongo in a drier savannah area ~350 km to the north with more seasonally-restricted transmission. Parasite DNA was sequenced and paired-end reads mapped to the P. falciparum reference genome. Results High coverage genome wide sequence data for 85 different clinical isolates enabled analysis of 121,712 single nucleotide polymorphisms (SNPs). The local populations had similar proportions of mixed genotype infections, similar SNP allele frequency distributions, and eleven chromosomal regions had elevated integrated haplotype scores (|iHS|) in both. A between-population Rsb metric comparing extended haplotype homozygosity indicated a stronger signal within Kintampo for one of these regions (on chromosome 14) and in Navrongo for two of these regions (on chromosomes 10 and 13). At least one gene in each of these identified regions is a potential target of locally varying selection. The candidates include genes involved in parasite development in mosquitoes, members of variant-expressed multigene families, and a leading vaccine-candidate target of immunity. Conclusions Against a background of very similar population structure and selection signatures in the P. falciparum populations of Ghana, three narrow genomic regions showed evidence indicating local differences in historical timing or intensity of selection. Sampling of closely situated populations across heterogeneous environments has potential to refine the mapping of important loci under temporally or spatially varying selection. Electronic supplementary material The online version of this article (doi:10.1186/s12864-015-1746-3) contains supplementary material, which is available to authorized users.

BACKGROUND: As countries make progress in malaria control, transmission may be reduced to such an extent that few cases occur, and identification of the remaining foci of transmission may require a combination of surveillance tools. The study explored the usefulness of parasite prevalence, seroprevalence and model-estimated seroconversion rates for detecting local differences in malaria transmission in a West African country. METHODS: Age-stratified cross-sectional surveys were conducted during the wet season in 2008 and the following dry season in 2009 in The Gambia. In each season, 20 village communities were sampled from six diverse areas throughout the country. A total of 7,586 participants were surveyed, 51% (3,870) during the wet season. Parasites were detected by thick film slide microscopy, and anti-MSP1-19 antibodies were detected by ELISA using eluted dried blood from filter papers. RESULTS: Overall parasite prevalence was 12.4% in the wet season and 2.2% in the dry season, with village-specific parasite prevalence ranging from 1.4 to 45.9% in the wet season and from 0.0 to 13.2% in the dry season. Prevalence was highest in the eastern part of the country. Serological indices also varied between villages, indicating local heterogeneity in transmission, and there was a high correlation between wet and dry season estimates across the villages. The overall prevalence of anti-MSP119 antibodies was similar in the wet (19.5%) and in the dry (19.6%) seasons. CONCLUSION: The study illustrates the utility of measuring both parasite prevalence and serological indices for monitoring local variation in malaria transmission, which are more informative than single measures as control intensifies and malaria declines. Measurements of seropositivity have the logistical advantage of being relative stable seasonally so that sampling at any time of year may be conducted.

Background Malaria parasite population genetic structure varies among areas of differing endemicity, but this has not been systematically studied across Plasmodium falciparum populations in Africa where most infections occur. Methods Ten polymorphic P. falciparum microsatellite loci were genotyped in 268 infections from eight locations in four West African countries (Republic of Guinea, Guinea Bissau, The Gambia and Senegal), spanning a highly endemic forested region in the south to a low endemic Sahelian region in the north. Analysis was performed on proportions of mixed genotype infections, genotypic diversity among isolates, multilocus standardized index of association, and inter-population differentiation. Results Each location had similar levels of pairwise genotypic diversity among isolates, although there were many more mixed parasite genotype infections in the south. Apart from a few isolates that were virtually identical, the multilocus index of association was not significant in any population. Genetic differentiation between populations was low (most pairwise F ST values Conclusions Although proportions of mixed genotype infections varied with endemicity as expected, population genetic structure was similar across the diverse sites. Very substantial reduction in transmission would be needed to cause fragmented or epidemic sub-structure in this region.

The transcription factor NFkappaB regulates inflammatory and other cellular responses. In non-stimulated cells, NFkappaB is linked to its inhibitor IkappaB, which plays a major role in controlling NFkappaB activity. Here, the gene promoter region of the major inducible IkappaB component (IkappaB-alpha) was studied to identify single nucleotide polymorphisms (SNPs), and to test if these are associated with risk of two diseases involving inflammation and fibrosis (trachoma and silicosis). Three SNPs were identified at positions -881, -826 and -297 relative to the transcription start site. The position -297 is close to two NFkappaB binding sites, kappaB2 and kappaB3, but the alleles were not associated with either disease. Alleles at positions -881 and -826 were in complete linkage disequilibrium with each other, and the rare haplotype was significantly less frequent among patients with trachoma compared to controls, although there was no difference in frequencies between silicosis patients and controls.

Background Health record-based observations from several parts of Africa indicate a major decline in malaria, but up-to-date information on parasite prevalence in West-Africa is sparse. This study aims to provide parasite prevalence data from three sites in the Gambia and Guinea Bissau, respectively, and compares the usefulness of PCR, rapid diagnostic tests (RDT), serology and slide-microscopy for surveillance. Methods Cross-sectional surveys in 12 villages at three rural sites were carried out in the Gambia and Guinea Bissau in January/February 2008, shortly following the annual transmission season. Results A surprisingly low microscopically detectable parasite prevalence was detected in the Gambia (Farafenni: 10.9%, CI95%: 8.7-13.1%; Basse: 9.0%, CI95%: 7.2-10.8%), and Guinea Bissau (Caio: 4%, CI95%: 2.6-5.4%), with low parasite densities (geometric mean: 104 parasites/μl, CI95%: 76-143/μl). In comparison, PCR detected a more than three times higher proportion of parasite carriers, indicating its usefulness to sensitively identify foci where malaria declines, whereas the RDT had very low sensitivity. Estimates of force of infection using age sero-conversion rates were equivalent to an EIR of approximately 1 infectious bite/person/year, significantly less than previous estimates. The sero-prevalence profiles suggest a gradual decline of malaria transmission, confirming their usefulness in providing information on longer term trends of transmission. A greater variability in parasite prevalence among villages within a site than between sites was observed with all methods. The fact that serology equally captured the inter-village variability, indicates that the observed heterogeneity represents a stable pattern. Conclusion PCR and serology may be used as complementary tools to survey malaria in areas of declining malaria prevalence such as the Gambia and Guinea Bissau.