Genome sequences of 247 Plasmodium falciparum isolates collected in The Gambia in 2008 and 2014 were analysed to identify changes possibly related to the scale-up of antimalarial interventions that occurred during this period. Overall, there were 15 regions across the genomes with signatures of positive selection. Five of these were sweeps around known drug resistance and antigenic loci. Signatures at antigenic loci such as thrombospodin related adhesive protein (Pftrap) were most frequent in eastern Gambia, where parasite prevalence and transmission remain high. There was a strong temporal differentiation at a non-synonymous SNP in a cysteine desulfarase (Pfnfs) involved in iron-sulphur complex biogenesis. During the 7-year period, the frequency of the lysine variant at codon 65 (Pfnfs-Q65K) increased by 22% (10% to 32%) in the Greater Banjul area. Between 2014 and 2015, the frequency of this variant increased by 6% (20% to 26%) in eastern Gambia. IC50 for lumefantrine was significantly higher in Pfnfs-65K isolates. This is probably the first evidence of directional selection on Pfnfs or linked loci by lumefantrine. Given the declining malaria transmission, the consequent loss of population immunity, and sustained drug pressure, it is important to monitor Gambian P. falciparum populations for further signs of adaptation.

Antimalarial interventions have yielded a significant decline in malaria prevalence in The Gambia, where artemether-lumefantrine (AL) has been used as a first-line antimalarial for a decade. Clinical Plasmodium falciparum isolates collected from 2012 to 2015 were analyzed ex vivo for antimalarial susceptibility and genotyped for drug resistance markers (pfcrt K76T, pfmdr1 codons 86, 184, and 1246, and pfk13) and microsatellite variation. Additionally, allele frequencies of single nucleotide polymorphisms (SNPs) from other drug resistance-associated genes were compared from genomic sequence data sets from 2008 (n = 79) and 2014 (n = 168). No artemisinin resistance-associated pfk13 mutation was found, and only 4% of the isolates tested in 2015 showed significant growth after exposure to dihydroartemisinin. Conversely, the 50% inhibitory concentrations (IC50s) of amodiaquine and lumefantrine increased within this period. pfcrt 76T and pfmdr1 184F mutants remained at a prevalence above 80%. pfcrt 76T was positively associated with higher IC50s to chloroquine. pfmdr1 NYD increased in frequency between 2012 and 2015 due to lumefantrine selection. The TNYD (pfcrt 76T and pfmdr1 NYD wild-type haplotype) also increased in frequency following AL implementation in 2008. These results suggest selection for pfcrt and pfmdr1 genotypes that enable tolerance to lumefantrine. Increased tolerance to lumefantrine calls for sustained chemotherapeutic monitoring in The Gambia to minimize complete artemisinin combination therapy (ACT) failure in the future.

Background In areas where Plasmodium falciparum malaria is seasonal, a dry season reservoir of blood-stage infection is essential for initiating transmission during the following wet season. Methods In The Gambia, a cohort of 42 individuals with qPCR positive P. falciparum infections at the end of the transmission season (December) were followed monthly until the end of the dry season (May) to evaluate infection persistence. The influence of human host and parasitological factors was investigated. Results A large proportion of individuals infected at the end of the wet season had detectable infections until the end of the dry season (40.0%; 16/40). At the start of the dry season, the majority of these persistent infections (82%) had parasite densities >10 p/µL compared to only 5.9% of short-lived infections. Persistent infections (59%) were also more likely to be multi-clonal than short-lived infections (5.9%) and were associated with individuals having higher levels of P. falciparum-specific antibodies (p = 0.02). Conclusion Asymptomatic persistent infections were multi-clonal with higher parasite densities at the beginning of the dry season. Screening and treating asymptomatic infections during the dry season may reduce the human reservoir of malaria responsible for initiating transmission in the wet season.

BackgroundIn areas where Plasmodium falciparum malaria is highly seasonal, a dry season reservoir of blood-stage infection is essential for initiating transmission during the following wet season, bridging transmission seasons several months apart. Understanding infections during the dry season could thus inform approaches for malaria control.MethodsIn The Gambia, a cohort of 42 individuals with qPCR positive P. falciparum infections at the end of the transmission season (December) were followed monthly until the end of the dry season (May) to evaluate the duration of detectable infections. The influence of human host (age, sex, haemoglobin concentration and genotype, and P. falciparum-specific antibodies), and parasitological (parasite density, gametocyte density and genotypic multiplicity of infection) factors was investigated.ResultsA large proportion of individuals infected at the end of the wet season had detectable infections until the end of the dry season (40.0%; 16/40), with the majority of these infections also harbouring gametocytes (81.3%; 13/16). 22 infections were classified as persistent (detectable for at least 3 months), 17 were classified as short-lived (undetectable within 2 months), and 3 were treated (due to symptoms). At the start of the dry season, the majority of persistent infections (82%; 18/22) had parasite densities >10 p/µL compared to only 5.9% (1/17) of short-lived infections. Persistent infections (59%; 13/22) were also more likely to be multi-clonal than short-lived infections (5.9%; 1/17), they were most common in 5 to 15 year old children (63%; 12/19), and were associated with individuals having higher levels of P. falciparum-specific antibodies (p = 0.058).ConclusionsAsymptomatic persistent dry season infections in The Gambia were multiclonal with higher parasite densities at the beginning of the dry season, mostly occurring in school age children and adults with higher P. falciparum-specific antibodies. Screening and treating asymptomatic, malaria-infected individuals during the dry season may reduce the human reservoir of malaria responsible initiating transmission in the wet-season.

BackgroundPrevious genome-wide analyses of single nucleotide variation in Plasmodium falciparum identified evidence of an extended haplotype region on chromosome 6 in West Africa, suggesting recent positive selection. Such a pattern is not seen in samples from East Africa or South East Asia, so it could be marking a selective process specific to West Africa. Analyses of the haplotype structure in samples taken at different times could give clues to possible causes of selection.MethodsThis study investigates chromosome 6 extended haplotypes in The Gambia by analysing alleles at multiple microsatellite loci using genome sequence data previously obtained from clinical isolates collected in 2008, followed by genotyping of 13 loci in 439 isolates from 1984, 1991, 2008 and 2014. Temporal changes in haplotype structure and frequencies were determined.ResultsA region of high linkage disequilibrium spanning over 170 kilobases (kb) was identified with both NGS and laboratory determined microsatellite alleles. Multiple long haplotypes were found in all temporal populations from The Gambia. Two of the haplotypes were detected in samples from 1984 and 1991. The frequency of long-range haplotypes increased in 2008 and 2014 populations. There was higher Fst between older and more recent populations at loci in proximity to genes involved in drug metabolism pathways.ConclusionsThe occurrence of several long haplotypes at intermediate frequencies suggests an unusual mode of selection in chromosome 6, possibly combined with recombination suppression on specific haplotypes. Such selection apparently occurred before the emergence of known anti-malarial drug resistance alleles, and could be due to effects of other drugs or unknown processes that have long been operating in this endemic region.Electronic supplementary materialThe online version of this article (doi:10.1186/s12936-016-1560-7) contains supplementary material, which is available to authorized users.