Isabel Gordo

Positive Epistasis Drives the Acquisition of Multidrug Resistance

The evolution of multiple antibiotic resistance is an increasing global problem. Resistance mutations are known to impair fitness, and the evolution of resistance to multiple drugs depends both on their costs individually and on how they interact—epistasis. Information on the level of epistasis between antibiotic resistance mutations is of key importance to understanding epistasis amongst deleterious alleles, a key theoretical question, and to improving public health measures. Here we show that in an antibiotic-free environment the cost of multiple resistance is smaller than expected, a signature of pervasive positive epistasis among alleles that confer resistance to antibiotics. Competition assays reveal that the cost of resistance to a given antibiotic is dependent on the presence of resistance alleles for other antibiotics. Surprisingly we find that a significant fraction of resistant mutations can be beneficial in certain resistant genetic backgrounds, that some double resistances entail no measurable cost, and that some allelic combinations are hotspots for rapid compensation. These results provide additional insight as to why multi-resistant bacteria are so prevalent and reveal an extra layer of complexity on epistatic patterns previously unrecognized, since it is hidden in genome-wide studies of genetic interactions using gene knockouts.

The evolution of a conjugative plasmid and its ability to increase bacterial fitness

Conjugative plasmids are extra-chromosomal DNA elements that are capable of horizontal transmission and are found in many natural isolated bacteria. Although plasmids may carry beneficial genes to their bacterial host, they may also cause a fitness cost. In this work, we studied the evolution of the R1 plasmid and we found that, in spite of the R1 plasmid conferring an initial cost to its host, after 420 generations the cost disappeared in all five independent evolution experiments. In fact, in two of these five experiments evolved conjugative plasmids actually conferred a fitness advantage to their hosts. Furthermore, the relative fitness of the ancestral clone bearing one of the evolved plasmids is significantly higher than both the plasmid-free ancestral cells and the evolved cells carrying the evolved plasmid. Given that the R1 plasmid may spread among different species of enterobacteria, we wondered what the effect of the evolved plasmid would be inside Salmonella enterica cells. We found that the evolved plasmid is also able to dramatically increase the relative fitness of these cells. Our results suggest that even if general usage of antibiotics is halted, conjugative plasmids that have been selected with antibiotics in previous years can still persist among bacterial populations or even invade new strains.

ADAPTIVE EVOLUTION OF ASEXUAL POPULATIONS UNDER MULLER'S RATCHET

Abstract We study the population genetics of adaptation in nonequilibrium haploid asexual populations. We find that the accumulation of deleterious mutations, due to the operation of Mullers ratchet, can considerably reduce the rate of fixation of advantageous alleles. Such reduction can be approximated reasonably well by a reduction in the effective population size. In the absence of Mullers ratchet, a beneficial mutation can only become fixed if it creates the best possible genotype; if Mullers ratchet operates, however, mutations initially arising in a nonoptimal genotype can also become fixed in the population, since the loss of the least‐loaded class implies that an initially nonoptimal background can become optimal. We show that, while the rate at which adaptive mutations become fixed is reduced, the rate of fixation of deleterious mutations due to the ratchet is not changed by the presence of beneficial mutations as long as the rate of their occurrence is low and the deleterious effects of mutations (sd) are higher than the beneficial effects (sa). When sa>sd, the advantage of a beneficial mutation can outweigh the deleterious effects of associated mutations. Under these conditions, a beneficial allele can drag to fixation deleterious mutations initially associated with it at a higher rate than in the absence of advantageous alleles. We propose analytical approximations for the rates of accumulation of deleterious and beneficial mutations. Furthermore, when allowing for the possible occurrence of interference between beneficial alleles, we find that the presence of deleterious mutations of either very weak or very strong effect can marginally increase the rate of accumulation of beneficial mutations over that observed in the absence of such deleterious mutations.

Genetic Recombination and Molecular Evolution

Reduced rates of genetic recombination are often associated with reduced genetic variability and levels of adaptation. Several different evolutionary processes, collectively known as Hill-Robertson (HR) effects, have been proposed as causes of these correlates of recombination. Here, we use DNA sequence polymorphism and divergence data from the noncrossing over dot chromosome of Drosophila to discriminate between two of the major forms of HR effects: selective sweeps and background selection. This chromosome shows reduced levels of silent variability and reduced effectiveness of selection. We show that neither model fits the data on variability. We propose that, in large genomic regions with restricted recombination, HR effects among nonsynonymous mutations undermine the effective strength of selection, so that their background selection effects are weakened. This modified model fits the data on variability and also explains why variability in very large nonrecombining genomes is not completely wiped out. We also show that HR effects of this type can produce an individual selection advantage to recombination, as well as greatly reduce the mean fitness of nonrecombining genomes and genomic regions.

Genetic linkage and molecular evolution

The different models make similar predictions which makes it difficult to ascribe an observation to just one process. Are there any observations that allow one to identify which process is operating? It turns out that the distribution of frequencies of neu-tral variants in a sample of sequences (the frequency spectrum) is model-dependent. A large distortion of the frequency spectrum towards rare variants is more likely with selective sweeps (Fig. 1Fig. 1). And when there is some recombination, so that initially rare variants are not necessarily swept to complete fixation, a transient sig-nature of a sweep is provided by the occurrence of an excess of ‘derived’ variants at a high frequency within a sample. (One can infer if a variant is derived or ancestral from the sequence of a closely related species.) Other features of genetic variation, such as patterns of linkage disequilibrium, may also be useful in testing alternative models. Perhaps the best way to quantify the relative importance of these processes is to get solid estimates of the rate at which deleterious mutations occur, and of the distribution of their effects on fitness. Although this is a simple question to ask, it is hard to answer. But given such information, and using the increasing amount of information on DNA sequence variation and evolution, one can perhaps try to answer an even harder question: what is the rate at which advantageous mutations occur, and what are their effects on fitness?Key references

Pervasive sign epistasis between conjugative plasmids and drug-resistance chromosomal mutations.

Multidrug-resistant bacteria arise mostly by the accumulation of plasmids and chromosomal mutations. Typically, these resistant determinants are costly to the bacterial cell. Yet, recently, it has been found that, in Escherichia coli bacterial cells, a mutation conferring resistance to an antibiotic can be advantageous to the bacterial cell if another antibiotic-resistance mutation is already present, a phenomenon called sign epistasis. Here we study the interaction between antibiotic-resistance chromosomal mutations and conjugative (i.e., self-transmissible) plasmids and find many cases of sign epistasis (40%)—including one of reciprocal sign epistasis where the strain carrying both resistance determinants is fitter than the two strains carrying only one of the determinants. This implies that the acquisition of an additional resistance plasmid or of a resistance mutation often increases the fitness of a bacterial strain already resistant to antibiotics. We further show that there is an overall antagonistic interaction between mutations and plasmids (52%). These results further complicate expectations of resistance reversal by interdiction of antibiotic use.

The first steps of adaptation of Escherichia coli to the gut are dominated by soft sweeps.

The accumulation of adaptive mutations is essential for survival in novel environments. However, in clonal populations with a high mutational supply, the power of natural selection is expected to be limited. This is due to clonal interference - the competition of clones carrying different beneficial mutations - which leads to the loss of many small effect mutations and fixation of large effect ones. If interference is abundant, then mechanisms for horizontal transfer of genes, which allow the immediate combination of beneficial alleles in a single background, are expected to evolve. However, the relevance of interference in natural complex environments, such as the gut, is poorly known. To address this issue, we have developed an experimental system which allows to uncover the nature of the adaptive process as Escherichia coli adapts to the mouse gut. This system shows the invasion of beneficial mutations in the bacterial populations and demonstrates the pervasiveness of clonal interference. The observed dynamics of change in frequency of beneficial mutations are consistent with soft sweeps, where different adaptive mutations with similar phenotypes, arise repeatedly on different haplotypes without reaching fixation. Despite the complexity of this ecosystem, the genetic basis of the adaptive mutations revealed a striking parallelism in independently evolving populations. This was mainly characterized by the insertion of transposable elements in both coding and regulatory regions of a few genes. Interestingly, in most populations we observed a complete phenotypic sweep without loss of genetic variation. The intense clonal interference during adaptation to the gut environment, here demonstrated, may be important for our understanding of the levels of strain diversity of E. coli inhabiting the human gut microbiota and of its recombination rate.

The speed of Muller's ratchet with background selection, and the degeneration of Y chromosomes

The rate of accumulation of deleterious mutations by Mullers ratchet is investigated in large asexual haploid populations, for a range of parameters with potential biological relevance. The rate of this process is studied by considering a very simple model in which mutations can have two types of effect: either strongly deleterious or mildly deleterious. It is shown that the rate of accumulation of mildly deleterious mutations can be greatly increased by the presence of strongly deleterious mutations, and that this can be predicted from the associated reduction in effective population size (the background selection effect). We also examine the rate of the ratchet when there are two classes of mutation of similar but unequal effects on fitness. The accuracy of analytical approximations for the rate of this process is analysed. Its possible role in causing the degeneration of Y and neo-Y chromosomes is discussed in the light of our present knowledge of deleterious mutation rates and selection coefficients.

Genome architecture is a selectable trait that can be maintained by antagonistic pleiotropy.

Chromosomal rearrangements are mutations contributing to both within and between species variation; however their contribution to fitness is yet to be measured. Here we show that chromosomal rearrangements are pervasive in natural isolates of Schizosaccharomyces pombe and contribute to reproductive isolation. To determine the fitness effects of chromosome structure, we constructed two inversions and eight translocations without changing the coding sequence. We show that chromosomal rearrangements contribute to both reproductive success in meiosis and growth rate in mitosis with a strong genotype by environment interaction. These changes are accompanied by alterations in gene expression. Strikingly, we find several examples leading to antagonistic pleiotropy. Even though chromosomal rearrangements may have a deleterious effect during sexual reproduction, some compensate with a strong growth advantage in mitosis. Our results constitute the first quantification of fitness effects caused by de novo mutations that result in chromosomal rearrangement variation and suggest a mechanism for their maintenance in natural populations.

Can intra-Y gene conversion oppose the degeneration of the human Y chromosome? A simulation study

The human Y is a genetically degenerate chromosome, which has lost about 97% of the genes originally present. Most of the remaining human Y genes are in large duplicated segments (ampliconic regions) undergoing intense Y–Y gene conversion. It has been suggested that Y–Y gene conversion may help these genes getting rid of deleterious mutations that would inactivate them otherwise. Here, we tested this idea by simulating the evolution of degenerating Y chromosomes with or without gene conversion using the most up-to-date population genetics parameters for humans. We followed the fate of a variant with Y–Y gene conversion in a population of Y chromosomes where Y–Y gene conversion is originally absent. We found that this variant gets fixed more frequently than the neutral expectation, which supports the idea that gene conversion is beneficial for a degenerating Y chromosome. Interestingly, a very high rate of gene conversion is needed for an effect of gene conversion to be observed. This suggests that high levels of Y-Y gene conversion observed in humans may have been selected to oppose the Y degeneration. We also studied with a similar approach the evolution of ampliconic regions on the Y chromosomes and found that the fixation of many copies at once is unlikely, which suggest these regions probably evolved gradually unless selection for increased dosage favored large-scale duplication events. Exploring the parameter space showed that Y–Y gene conversion may be beneficial in most mammalian species, which is consistent with recent data in chimpanzees and mice.