The results we present here on malaria population biology are important in that they confirm with significantly more data previous observations about the distribution of diversity in the P. falciparum
parasite population, highlight the influence of natural selection on P. falciparum
genomic diversity and divergence, and provide strong evidence that larger datasets will prove very useful in learning more about the biology of the parasite and the manifestation of disease. For example, though previous multilocus genotyping efforts in this parasite succeeded in documenting heterogeneous diversity levels and genetic differentiation among populations, the scale of the present analysis has enabled us to statistically distinguish the pattern of differentiation at nonsynonymous SNPs potentially subject to natural selection and synonymous or silent SNPs that are presumably closer to selective neutrality, but otherwise subject to similar effects from demographic effects and nearby selective sweeps. We observed higher FST
values for nonsynonymous SNPs than for silent SNPs in all population comparisons except Africa versus Americas (which exhibited highly similar values for both classes), which suggests a role for natural selection in the ongoing genetic differentiation of malaria populations in disparate geographic locales. Though a previous investigation observed a similar contrast in divergence between ten nonsynonymous SNPs at loci known to be associated with anti-malarial drug resistance and ten silent SNPs at housekeeping or unknown loci [23
], we have demonstrated that nonsynonymous variation exhibits a distinct evolutionary profile across the genome, and not only at loci known to be subject to drug-mediated selection.
Both negative (purifying), and positive (directional/balancing) selection can increase population divergence if the efficacy of the selection differs geographically [24
]. A recent study of human SNP data found that nonsynonymous SNPs were more likely than synonymous SNPs to exhibit extremes of both low divergence (attributed by the authors to purifying selection) and high divergence (attributed to positive selection) [25
]. Our data show only half of this pattern, an overall trend for nonsynonymous SNPs to have higher divergence than silent ones. This pattern suggests that either localized positive selection or strong differences in selective constraints play a substantial role in shaping population differentiation in P. falciparum
(the failure to detect the clearest signal of purifying selection, an excess of low-FST
nonsynonymous SNPs, may simply result from small sample size).
Even if negative selection plays an important role in nonsynonymous SNP divergence among P. falciparum populations, there are numerous potential population-specific environmental factors that could induce differentiation by positive selection. For example, population differentiation at nonsynonymous SNPs could result in variation among human host populations in HLA allelic composition, different Anopheles mosquito vectors, and/or heterogeneous application of anti-malarial therapeutics. Alternatively, some of the variant nonsynonymous SNPs may represent positively selected mutations that have arisen due to selection pressures that are present in both populations, but which have been prevented from spreading between the populations due to geographic barriers to gene flow. Deeper probing of these questions with expanded sample sets and denser SNP assays should illuminate whether these observed patterns of divergence and polymorphism are due to negative, positive, or balancing selection pressures.
Nonsynonymous SNPs that vary widely in frequency among P. falciparum populations warrant deeper follow-up investigations, as many may represent functional variations with important implications for public health or understanding of basic malaria biology. Nonsynonymous SNPs that differ even marginally in frequency among populations may be subject to selection (Figure ). It should be noted that while the SNPs selected for inclusion in our pilot array are biased for a number of reasons, they are not expected to be enriched for variants subject to selection. Indeed, nonsynonymous SNPs in highly polymorphic subtelomeric gene families are known to be subject to strong immune selection, but are virtually absent among the SNPs on our array due to difficulties in recognizing and genotyping SNPs that are in close physical proximity. In all, only 12 of our SNPs were in clearly identified antigenic genes (as defined by Gene Ontology category).
The SNPs we profiled, consequently, may be expected to yield a conservative estimate of the degree to which segregating genetic variation in P. falciparum
is subject to population-specific positive or negative natural selection. This broad survey of genetic diversity in P. falciparum
stands in contrast to most previous analyses of variability in the parasite, many of which focused on antigenic loci such as AMA-1
] or MSP-1
] that are known to be subject to strong positive selection. While these population genetic analyses of antigens yield valuable information about the loci in question, the influence of strong host-mediated selective pressure erases typical patterns of genetic diversity and LD in the region of these genes, making difficult any accurate inferences about demographic history or population structure from the data. The very short range of detectable LD on chromosome 7 makes it likely that the profile of diversity we have observed is not dominated by one or more linked loci subject to strong selection (such as pfcrt
), but instead that it reflects many independent, relatively unbiased local profiles of the interplay of natural selection and genetic drift.