Background: The study of reverse evolution from resistant to susceptible phenotypes can reveal constraints on biological evolution, a topic for which evolutionary theory has relatively few general principles. The public health catastrophe of antimicrobial resistance in malaria has brought these constraints on evolution into a practical realm, with one proposed solution: withdrawing anti-malarial medication use in high resistance settings, built on the assumption that reverse evolution occurs readily enough that populations of pathogens may revert to their susceptible states. While past studies have suggested limits to reverse evolution, there have been few attempts to properly dissect its mechanistic constraints. Methods: Growth rates were determined from empirical data on the growth and resistance from a set of combinatorially complete set of mutants of a resistance protein (dihydrofolate reductase) in Plasmodium vivax, to construct reverse evolution trajectories. The fitness effects of individual mutations were calculated as a function of drug environment, revealing the magnitude of epistatic interactions between mutations and genetic backgrounds. Evolution across the landscape was simulated in two settings: starting from the population fixed for the quadruple mutant, and from a polymorphic population evenly distributed between double mutants. Results: A single mutation of large effect (S117N) serves as a pivot point for evolution to high resistance regions of the landscape. Through epistatic interactions with other mutations, this pivot creates an epistatic ratchet against reverse evolution towards the wild type ancestor, even in environments where the wild type is the most fit of all genotypes. This pivot mutation underlies the directional bias in evolution across the landscape, where evolution towards the ancestor is precluded across all examined drug concentrations from various starting points in the landscape. Conclusions: The presence of pivot mutations can dictate dynamics of evolution across adaptive landscape through epistatic interactions within a protein, leaving a population trapped on local fitness peaks in an adaptive landscape, unable to locate ancestral genotypes. This irreversibility suggests that the structure of an adaptive landscape for a resistance protein should be understood before considering resistance management strategies. This proposed mechanism for constraints on reverse evolution corroborates evidence from the field indicating that phenotypic reversal often occurs via compensatory mutation at sites independent of those associated with the forward evolution of resistance. Because of this, molecular methods that identify resistance patterns via single SNPs in resistance-associated markers might be missing signals for resistance and compensatory mutation throughout the genome. In these settings, whole genome sequencing efforts should be used to identify resistance patterns, and will likely reveal a more complicated genomic signature for resistance and susceptibility, especially in settings where anti-malarial medications have been used intermittently. Lastly, the findings suggest that, given their role in dictating the dynamics of evolution across the landscape, pivot mutations might serve as future targets for therapy. Electronic supplementary material The online version of this article (doi:10.1186/s12936-016-1090-3) contains supplementary material, which is available to authorized users.
The adaptive landscape analogy has found practical use in recent years, as many have explored how their understanding can inform therapeutic strategies that subvert the evolution of drug resistance. A major barrier to applications of these concepts is a lack of detail concerning how the environment affects adaptive landscape topography, and consequently, the outcome of drug treatment. Here we combine empirical data, evolutionary theory, and computer simulations towards dissecting adaptive landscape by environment interactions for the evolution of drug resistance in two dimensions—drug concentration and drug type. We do so by studying the resistance mediated by Plasmodium falciparum dihydrofolate reductase (DHFR) to two related inhibitors—pyrimethamine and cycloguanil—across a breadth of drug concentrations. We first examine whether the adaptive landscapes for the two drugs are consistent with common definitions of cross-resistance. We then reconstruct all accessible pathways across the landscape, observing how their structure changes with drug environment. We offer a mechanism for non-linearity in the topography of accessible pathways by calculating of the interaction between mutation effects and drug environment, which reveals rampant patterns of epistasis. We then simulate evolution in several different drug environments to observe how these individual mutation effects (and patterns of epistasis) influence paths taken at evolutionary “forks in the road” that dictate adaptive dynamics in silico. In doing so, we reveal how classic metrics like the IC50 and minimal inhibitory concentration (MIC) are dubious proxies for understanding how evolution will occur across drug environments. We also consider how the findings reveal ambiguities in the cross-resistance concept, as subtle differences in adaptive landscape topography between otherwise equivalent drugs can drive drastically different evolutionary outcomes. Summarizing, we discuss the results with regards to their basic contribution to the study of empirical adaptive landscapes, and in terms of how they inform new models for the evolution of drug resistance.
Fitness landscapes of drug resistance constitute powerful tools to elucidate mutational pathways of antibiotic escape. Here, we developed a predictive biophysics-based fitness landscape of trimethoprim (TMP) resistance for Escherichia coli dihydrofolate reductase (DHFR). We investigated the activity, binding, folding stability, and intracellular abundance for a complete set of combinatorial DHFR mutants made out of three key resistance mutations and extended this analysis to DHFR originated from Chlamydia muridarum and Listeria grayi. We found that the acquisition of TMP resistance via decreased drug affinity is limited by a trade-off in catalytic efficiency. Protein stability is concurrently affected by the resistant mutants, which precludes a precise description of fitness from a single molecular trait. Application of the kinetic flux theory provided an accurate model to predict resistance phenotypes (IC50) quantitatively from a unique combination of the in vitro protein molecular properties. Further, we found that a controlled modulation of the GroEL/ES chaperonins and Lon protease levels affects the intracellular steady-state concentration of DHFR in a mutation-specific manner, whereas IC50 is changed proportionally, as indeed predicted by the model. This unveils a molecular rationale for the pleiotropic role of the protein quality control machinery on the evolution of antibiotic resistance, which, as we illustrate here, may drastically confound the evolutionary outcome. These results provide a comprehensive quantitative genotype–phenotype map for the essential enzyme that serves as an important target of antibiotic and anticancer therapies.
Crosses between closely related animal species often result in male hybrids that are sterile, and the molecular and functional basis of genetic factors for hybrid male sterility is of great interest. Here, we report a molecular and functional analysis of HMS1, a region of 9.2 kb in chromosome 3 of Drosophila mauritiana, which results in virtually complete hybrid male sterility when homozygous in the genetic background of sibling species Drosophila simulans. The HMS1 region contains two strong candidate genes for the genetic incompatibility, agt and Taf1. Both encode unrelated DNA-binding proteins, agt for an alkyl-cysteine-S-alkyltransferase and Taf1 for a subunit of transcription factor TFIID that serves as a multifunctional transcriptional regulator. The contribution of each gene to hybrid male sterility was assessed by means of germ-line transformation, with constructs containing complete agt and Taf1 genomic sequences as well as various chimeric constructs. Both agt and Taf1 contribute about equally to HMS1 hybrid male sterility. Transgenes containing either locus rescue sterility in about one-half of the males, and among fertile males the number of offspring is in the normal range. This finding suggests compensatory proliferation of the rescued, nondysfunctional germ cells. Results with chimeric transgenes imply that the hybrid incompatibilities result from interactions among nucleotide differences residing along both agt and Taf1. Our results challenge a number of preliminary generalizations about the molecular and functional basis of hybrid male sterility, and strongly reinforce the role of DNA-binding proteins as a class of genes contributing to the maintenance of postzygotic reproductive isolation.
Genes encode components of coevolved and interconnected networks. The effect of genotype on phenotype therefore depends on genotypic context through gene interactions known as epistasis. Epistasis is important in predicting phenotype from genotype for an individual. It is also examined in population studies to identify genetic risk factors in complex traits and to predict evolution under selection. Paradoxically, the effects of genotypic context in individuals and populations are distinct and sometimes contradictory. We argue that predicting genotype from phenotype for individuals based on population studies is difficult and, especially in human genetics, likely to result in underestimating the effects of genotypic context.
Plasmodium falciparum malaria remains a devastating public health problem. Recent discoveries have shed light on the origin and evolution of Plasmodium parasites and their interactions with their vertebrate and mosquito hosts. P. falciparum malaria originated in Africa from a single horizontal transfer between an infected gorilla and a human, and became global as the result of human migration. Today, P. falciparum malaria is transmitted worldwide by more than 70 different anopheline mosquito species. Recent studies indicate that the mosquito immune system can be a barrier to malaria transmission and that the P. falciparum Pfs47 gene allows the parasite to evade mosquito immune detection. Here, we review the origin and globalization of P. falciparum and integrate this history with analysis of the biology, evolution, and dispersal of the main mosquito vectors. This new perspective broadens our understanding of P. falciparum population structure and the dispersal of important parasite genetic traits.
Background: Encouraging advances in the control of Plasmodium falciparum malaria have been observed across much of Africa in the past decade. However, regions of high relative prevalence and transmission that remain unaddressed or unrecognized provide a threat to this progress. Difficulties in identifying such localized hotspots include inadequate surveillance, especially in remote regions, and the cost and labor needed to produce direct estimates of transmission. Genetic data can provide a much-needed alternative to such empirical estimates, as the pattern of genetic variation within malaria parasite populations is indicative of the level of local transmission. Here, genetic data were used to provide the first empirical estimates of P. falciparum malaria prevalence and transmission dynamics for the rural, remote Makira region of northeastern Madagascar. Methods: Longitudinal surveys of a cohort of 698 total individuals (both sexes, 0–74 years of age) were performed in two communities bordering the Makira Natural Park protected area. Rapid diagnostic tests, with confirmation by molecular methods, were used to estimate P. falciparum prevalence at three seasonal time points separated by 4-month intervals. Genomic loci in a panel of polymorphic, putatively neutral markers were genotyped for 94 P. falciparum infections and used to characterize genetic parameters known to correlate with transmission levels. Results: Overall, 27.8% of individuals tested positive for P. falciparum over the 10-month course of the study, a rate approximately sevenfold higher than the countrywide average for Madagascar. Among those P. falciparum infections, a high level of genotypic diversity and a high frequency of polygenomic infections (68.1%) were observed, providing a pattern consistent with high and stable transmission. Conclusions: Prevalence and genetic diversity data indicate that the Makira region is a hotspot of P. falciparum transmission in Madagascar. This suggests that the area should be highlighted for future interventions and that additional areas of high transmission may be present in ecologically similar regions nearby. Electronic supplementary material The online version of this article (doi:10.1186/s12936-016-1644-4) contains supplementary material, which is available to authorized users.