Multiple origins and regional dispersal of resistant dhps in African Plasmodium falciparum malaria

Abstract

Background: Although the molecular basis of resistance to a number of common antimalarial drugs is well known, a geographic description of the emergence and dispersal of resistance mutations across Africa has not been attempted. To that end we have characterised the evolutionary origins of antifolate resistance mutations in the dihydropteroate synthase (dhps) gene and mapped their contemporary distribution.

Methods and findings: We used microsatellite polymorphism flanking the dhps gene to determine which resistance alleles shared common ancestry and found five major lineages each of which had a unique geographical distribution. The extent to which allelic lineages were shared among 20 African Plasmodium falciparum populations revealed five major geographical groupings. Resistance lineages were common to all sites within these regions. The most marked differentiation was between east and west African P. falciparum, in which resistance alleles were not only of different ancestry but also carried different resistance mutations.

Conclusions: Resistant dhps has emerged independently in multiple sites in Africa during the past 10-20 years. Our data show the molecular basis of resistance differs between east and west Africa, which is likely to translate into differing antifolate sensitivity. We have also demonstrated that the dispersal patterns of resistance lineages give unique insights into recent parasite migration patterns.

Figure 1. Map of the countries of Africa included in this study.

Figure 2. The distribution of the major dhps alleles across sub-Saharan Africa.

Resistant alleles; the upper map shows the relative proportions of the three major resistance alleles, SGK, AGK, and SGE. Wild-type alleles; the lower map shows the ratio of SAK and AAK alleles among wild-type dhps alleles. In both cases the diameter of the pie is proportional to the combined frequencies of the alleles represented in the total population.

Figure 3. Microsatellite diversity around the wild-type (SAK and AAK) and the resistant (AGK, SGK, and SGE) alleles.

The expected heterozygosity (H _e) at flanking loci 0.8 kb, 4.3 kb, and 7.7 kb from the dhps gene was calculated for each geographical site (provided the number of observation ≥10), and box plots show the median, interquartile ranges, and the upper and lower extremes of the distribution of H _e values among geographical sites. Where there are statistical outliers, these are indicated by small squares.

Figure 4. Microsatellite polymorphism flanking wild-type and resistant dhps alleles.

In the bar graphs all the microsatellite haplotypes observed have been ranked first according to allele size at locus 0.8 kb and then by allele size at locus 4.3 kb along a common x-axis. The association of specific microsatellite haplotypes with different dhps alleles is apparent from the frequencies of each haplotype shown in the individual charts. (A) haplotypes linked to SAK AAK wild-type alleles, (B) haplotypes linked to AGK SGK single-mutant alleles, and (C) haplotypes linked to SGE double-mutant alleles.

Figure 5. The African distribution of dhps resistance lineages.

The distribution of the five major lineages among the geographic sites is indicated in the map. Resistance alleles whose flanking microsatellite haplotypes did not conform to a defined major lineage are shown in grey. Sharing of resistance allele lineages among the African populations is shown in a cladogram based on pairwise comparison of allele sharing (D _PS), which includes all the flanking haplotypes identified. Closely related populations cluster in large geographic regions that supercede national boundaries.