The Plasmodium falciparum parasite's ability to adapt to environmental pressures, such as the human immune system and antimalarial drugs, makes malaria an enduring burden to public health. Understanding the genetic basis of these adaptations is critical to intervening successfully against malaria. To that end, we created a high-density genotyping array that assays over 17,000 single nucleotide polymorphisms (∼1 SNP/kb), and applied it to 57 culture-adapted parasites from three continents. We characterized genome-wide genetic diversity within and between populations and identified numerous loci with signals of natural selection, suggesting their role in recent adaptation. In addition, we performed a genome-wide association study (GWAS), searching for loci correlated with resistance to thirteen antimalarials; we detected both known and novel resistance loci, including a new halofantrine resistance locus, PF10_0355. Through functional testing we demonstrated that PF10_0355 overexpression decreases sensitivity to halofantrine, mefloquine, and lumefantrine, but not to structurally unrelated antimalarials, and that increased gene copy number mediates resistance. Our GWAS and follow-on functional validation demonstrate the potential of genome-wide studies to elucidate functionally important loci in the malaria parasite genome.

Author Summary

Malaria infection with the human pathogen Plasmodium falciparum results in almost a million deaths each year, mostly in African children. Efforts to eliminate malaria are underway, but the parasite is adept at eluding both the human immune response and antimalarial treatments. Thus, it is important to understand how the parasite becomes resistant to drugs and to develop strategies to overcome resistance mechanisms. Toward this end, we used population genetic strategies to identify genetic loci that contribute to parasite adaptation and to identify candidate genes involved in drug resistance. We examined over 17,000 genetic variants across the parasite genome in over 50 strains in which we also measured responses to many known antimalarial compounds. We found a number of genetic loci showing signs of recent natural selection and a number of loci potentially involved in modulating the parasite's response to drugs. We further demonstrated that one of the novel candidate genes (PF10_0355) modulates resistance to the antimalarial compounds halofantrine, mefloquine, and lumefantrine. Overall, this study confirms that we can use genome-wide approaches to identify clinically relevant genes and demonstrates through functional testing that at least one of these candidate genes is indeed involved in antimalarial drug resistance.

Allpaths2, a method for accurately assembling small genomes with high continuity using short paired reads.

We demonstrate that genome sequences approaching finished quality can be generated from short paired reads. Using 36 base (fragment) and 26 base (jumping) reads from five microbial genomes of varied GC composition and sizes up to 40 Mb, ALLPATHS2 generated assemblies with long, accurate contigs and scaffolds. Velvet and EULER-SR were less accurate. For example, for Escherichia coli, the fraction of 10-kb stretches that were perfect was 99.8% (ALLPATHS2), 68.7% (Velvet), and 42.1% (EULER-SR).