Person:

Rice, Daniel

Background: Drug resistance remains a major public health challenge for malaria treatment and eradication. Individual loci associated with drug resistance to many antimalarials have been identified, but their epistasis with other resistance mechanisms has not yet been elucidated. Results: We previously described two mutations in the cytoplasmic prolyl-tRNA synthetase (cPRS) gene that confer resistance to halofuginone. We describe here the evolutionary trajectory of halofuginone resistance of two independent drug resistance selections in Plasmodium falciparum. Using this novel methodology, we discover an unexpected non-genetic drug resistance mechanism that P. falciparum utilizes before genetic modification of the cPRS. P. falciparum first upregulates its proline amino acid homeostasis in response to halofuginone pressure. We show that this non-genetic adaptation to halofuginone is not likely mediated by differential RNA expression and precedes mutation or amplification of the cPRS gene. By tracking the evolution of the two drug resistance selections with whole genome sequencing, we further demonstrate that the cPRS locus accounts for the majority of genetic adaptation to halofuginone in P. falciparum. We further validate that copy-number variations at the cPRS locus also contribute to halofuginone resistance. Conclusions: We provide a three-step model for multi-locus evolution of halofuginone drug resistance in P. falciparum. Informed by genomic approaches, our results provide the first comprehensive view of the evolutionary trajectory malaria parasites take to achieve drug resistance. Our understanding of the multiple genetic and non-genetic mechanisms of drug resistance informs how we will design and pair future anti-malarials for clinical use. Electronic supplementary material The online version of this article (doi:10.1186/s13059-014-0511-2) contains supplementary material, which is available to authorized users.

The dynamics of adaptation determines which mutations fix in a population, and hence how reproducible evolution will be. This is central to understanding the spectra of mutations recovered in evolution of antibiotic resistance1, the response of pathogens to immune selection2,3, and the dynamics of cancer progression4,5. In laboratory evolution experiments, demonstrably beneficial mutations are found repeatedly6–8, but are often accompanied by other mutations with no obvious benefit. Here we use whole-genome whole-population sequencing to examine the dynamics of genome sequence evolution at high temporal resolution in 40 replicate Saccharomyces cerevisiae populations growing in rich medium for 1,000 generations. We find pervasive genetic hitchhiking: multiple mutations arise and move synchronously through the population as mutational “cohorts.” Multiple clonal cohorts are often present simultaneously, competing with each other in the same population. Our results show that patterns of sequence evolution are driven by a balance between these chance effects of hitchhiking and interference, which increase stochastic variation in evolutionary outcomes, and the deterministic action of selection on individual mutations, which favors parallel evolutionary solutions in replicate populations.

Sex and recombination are pervasive throughout nature despite their substantial costs1. Understanding the evolutionary forces that maintain these phenomena is a central challenge in biology2,3. One longstanding hypothesis argues that sex is beneficial because recombination speeds adaptation4. Theory has proposed a number of distinct population genetic mechanisms that could underlie this advantage. For example, sex can promote the fixation of beneficial mutations either by alleviating interference competition (the Fisher-Muller effect)5,6 or by separating them from deleterious load (the ruby in the rubbish effect)7,8. Previous experiments confirm that sex can increase the rate of adaptation9–17, but these studies did not observe the evolutionary dynamics that drive this effect at the genomic level. Here, we present the first comparison between the sequence-level dynamics of adaptation in experimental sexual and asexual populations, which allows us to identify the specific mechanisms by which sex speeds adaptation. We find that sex alters the molecular signatures of evolution by changing the spectrum of mutations that fix, and confirm theoretical predictions that it does so by alleviating clonal interference. We also show that substantially deleterious mutations hitchhike to fixation in adapting asexual populations. In contrast, recombination prevents such mutations from fixing. Our results demonstrate that sex both speeds adaptation and alters its molecular signature by allowing natural selection to more efficiently sort beneficial from deleterious mutations.