Mutational neighbourhood and mutation supply rate constrain adaptation in Pseudomonas aeruginosa

Understanding adaptation by natural selection requires understanding the genetic factors that determine which beneficial mutations are available for selection. Here, using experimental evolution of rifampicin-resistant Pseudomonas aeruginosa, we show that different genotypes vary in their capacity for adaptation to the cost of antibiotic resistance. We then use sequence data to show that the beneficial mutations associated with fitness recovery were specific to particular genetic backgrounds, suggesting that genotypes had access to different sets of beneficial mutations. When we manipulated the supply rate of beneficial mutations, by altering effective population size during evolution, we found that it constrained adaptation in some selection lines by restricting access to rare beneficial mutations, but that the effect varied among the genotypes in our experiment. These results suggest that mutational neighbourhood varies even among genotypes that differ by a single amino acid change, and this determines their capacity for adaptation as well as the influence of population biology processes that alter mutation supply rate.     

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.     

Effects of genotype on rates of substitution during experimental evolution

Rates of molecular evolution may vary widely between populations, yet the causes of this variation are still incompletely understood. Genetic differences between populations may make an important contribution to variation in rates of evolution, owing to differences in fitness, population size, mutation rates, or in the distribution of fitness effects (DFEs) of available beneficial mutations. By whole genome sequencing of Escherichia coli populations experimentally evolved in the presence of a quinolone antibiotic, we found that rates of substitution varied by genotype, with evidence for a contribution from a genotype's starting fitness. Subsequent targeted sequencing showed that genotypes with high average substitution rates were more likely to undergo the simultaneous fixation of several mutations, consistent with theoretical models of multiple mutation dynamics. Moreover, patterns of substitution were indicative of epistatic relationships between known resistance mutations.     

Relative effects of segregation and recombination on the evolution of sex in finite diploid populations

The mechanism of reproducing more viable offspring in response to selection is a major factor influencing the advantages of sex. In diploids, sexual reproduction combines genotype by recombination and segregation. Theoretical studies of sexual reproduction have investigated the advantage of recombination in haploids. However, the potential advantage of segregation in diploids is less studied. This study aimed to quantify the relative contribution of recombination and segregation to the evolution of sex in finite diploids by using multilocus simulations. The mean fitness of a sexually or asexually reproduced population was calculated to describe the long-term effects of sex. The evolutionary fate of a sex or recombination modifier was also monitored to investigate the short-term effects of sex. Two different scenarios of mutations were considered: (1) only deleterious mutations were present and (2) a combination of deleterious and beneficial mutations. Results showed that the combined effects of segregation and recombination strongly contributed to the evolution of sex in diploids. If deleterious mutations were only present, segregation efficiently slowed down the speed of Muller''s ratchet. As the recombination level was increased, the accumulation of deleterious mutations was totally inhibited and recombination substantially contributed to the evolution of sex. The presence of beneficial mutations evidently increased the fixation rate of a recombination modifier. We also observed that the twofold cost of sex was easily to overcome in diploids if a sex modifier caused a moderate frequency of sex.     

The character or the variation: the genetic analysis of the insecticide-resistance phenotype

In this critique it is argued that the genetic basis of the evolution of resistance is dependent on how the phenotypic, and underlying genotypic, variation is channelled during a selective response. A polygenic response is preferentially favoured if selection acts within the phenotypic distribution of susceptibles; a monogenic response is predicted if selection screens rare mutations with phenotypes outside that susceptible distribution. The relevance of this model to the method of genetic analysis, the prediction of resistance mechanisms to novel insecticides, the generation of resistant beneficial insects and the development of the most effective resistance and integrated pest management programmes is discussed.     

Genomic Analysis Reveals Distinct Concentration-Dependent Evolutionary Trajectories for Antibiotic Resistance in Escherichia coli

Evolution of bacteria under sublethal concentrations of antibiotics represents a trade-off between growth and resistance to the antibiotic. To understand this trade-off, we performed in vitro evolution of laboratory Escherichia coli under sublethal concentrations of the aminoglycoside kanamycin over short time durations. We report that fixation of less costly kanamycin-resistant mutants occurred earlier in populations growing at lower sublethal concentration of the antibiotic, compared with those growing at higher sublethal concentrations; in the latter, resistant mutants with a significant growth defect persisted longer. Using deep sequencing, we identified kanamycin resistance-conferring mutations, which were costly or not in terms of growth in the absence of the antibiotic. Multiple mutations in the C-terminal end of domain IV of the translation elongation factor EF-G provided low-cost resistance to kanamycin. Despite targeting the same or adjacent residues of the protein, these mutants differed from each other in the levels of resistance they provided. Analysis of one of these mutations showed that it has little defect in growth or in synthesis of green fluorescent protein (GFP) from an inducible plasmid in the absence of the antibiotic. A second class of mutations, recovered only during evolution in higher sublethal concentrations of the antibiotic, deleted the C-terminal end of the ATP synthase shaft. This mutation confers basal-level resistance to kanamycin while showing a strong growth defect in the absence of the antibiotic. In conclusion, the early dynamics of the development of resistance to an aminoglycoside antibiotic is dependent on the levels of stress (concentration) imposed by the antibiotic, with the evolution of less costly variants only a matter of time.     

The cost of multiple drug resistance in Pseudomonas aeruginosa

The spread of bacterial antibiotic resistance mutations is thought to be constrained by their pleiotropic fitness costs. Here we investigate the fitness costs of resistance in the context of the evolution of multiple drug resistance (MDR), by measuring the cost of acquiring streptomycin resistance mutations (StrepR) in independent strains of the bacterium Pseudomonas aeruginosa carrying different rifampicin resistance (RifR) mutations. In the absence of antibiotics, StrepR mutations are associated with similar fitness costs in different RifR genetic backgrounds. The cost of StrepR mutations is greater in a rifampicin‐sensitive (RifS) background, directly demonstrating antagonistic epistasis between resistance mutations. In the presence of rifampicin, StrepR mutations have contrasting effects in different RifR backgrounds: StrepR mutations have no detectable costs in some RifR backgrounds and massive fitness costs in others. Our results clearly demonstrate the importance of epistasis and genotype‐by‐environment interactions for the evolution of MDR.     

The impact of transposable elements on eukaryotic genomes: From genome size increase to genetic adaptation to stressful environments

Transposable elements (TEs) are present in roughly all genomes. These mobile DNA sequences are able to invade genomes and their impact on genome evolution is substantial. The mobility of TEs can induce the appearance of deleterious mutations, gene disruption and chromosome rearrangements, but transposition activity also has positive aspects and the mutational activities of TEs contribute to the genetic diversity of organisms. This short review aims to give a brief overview of the impact TEs may have on animal and plant genome structure and expression, and the relationship between TEs and the stress response of organisms, including insecticide resistance.     

AMELIORATION OF THE DELETERIOUS PLEIOTROPIC EFFECTS OF AN ADAPTIVE MUTATION IN BACILLUS SUBTILIS

The deleterious pleiotropic effects of an adaptive mutation may be ameliorated by one of two modes of evolution: (1) by replacement, in which an adaptive mutation with harmful pleiotropic effects is replaced by one that confers an equal benefit but at less cost; or (2) by compensatory evolution, in which natural selection favors modifiers at other loci that compensate for the deleterious effects of the mutant allele. In this study, we have measured the potential of these two modes of evolution to ameliorate the deleterious pleiotropic effects of resistance to the antibiotic rifampicin in the soil bacterium Bacillus subtilis. One approach was to measure the fitness cost of a series of spontaneous rifampicin-resistance mutations from each of several strains. The potential for amelioration by the replacement mode was estimated by the variation in fitness cost among the mutants of a single strain. Another approach was to introduce a series of different rifampicin-resistance alleles into a diversity of strains, and to measure the fitness cost of rifampicin resistance for each allele-by-strain combination. The potential for amelioration by the replacement mode was estimated by the variation in fitness costs among rifampicin-resistance alleles; the potential for compensatory evolution was estimated by variation in the fitness cost of rifampicin resistance among strains. This study has shown that the cost of rifampicin resistance may be ameliorated by both the compensatory and replacement modes.     

The advantages and disadvantages of horizontal gene transfer and the emergence of the first species

Background  

Horizontal Gene Transfer (HGT) is beneficial to a cell if the acquired gene confers a useful function, but is detrimental if the gene has no function, if it is incompatible with existing genes, or if it is a selfishly replicating mobile element. If the balance of these effects is beneficial on average, we would expect cells to evolve high rates of acceptance of horizontally transferred genes, whereas if it is detrimental, cells should reduce the rate of HGT as far as possible. It has been proposed that the rate of HGT was very high in the early stages of prokaryotic evolution, and hence there were no separate lineages of organisms. Only when the HGT rate began to fall, would lineages begin to emerge with their own distinct sets of genes. Evolution would then become more tree-like. This phenomenon has been called the Darwinian Threshold.     

Characterization of mutations in the PAS domain of the EvgS sensor kinase selected by laboratory evolution for acid resistance in Escherichia coli

Laboratory‐based evolution and whole‐genome sequencing can link genotype and phenotype. We used evolution of acid resistance in exponential phase Escherichia coli to study resistance to a lethal stress. Iterative selection at pH 2.5 generated five populations that were resistant to low pH in early exponential phase. Genome sequencing revealed multiple mutations, but the only gene mutated in all strains was evgS, part of a two‐component system that has already been implicated in acid resistance. All these mutations were in the cytoplasmic PAS domain of EvgS, and were shown to be solely responsible for the resistant phenotype, causing strong upregulation at neutral pH of genes normally induced by low pH. Resistance to pH 2.5 in these strains did not require the transporter GadC, or the sigma factor RpoS. We found that EvgS‐dependent constitutive acid resistance to pH 2.5 was retained in the absence of the regulators GadE or YdeO, but was lost if the oxidoreductase YdeP was also absent. A deletion in the periplasmic domain of EvgS abolished the response to low pH, but not the activity of the constitutive mutants. On the basis of these results we propose a model for how EvgS may become activated by low pH.     

环境胁迫诱导的细胞适应性突变

细胞具有普遍的突变和进化能力, 如病原菌的抗药性、工业菌株的适应性和人体细胞的癌变等, 但是细胞的适应性突变是如何产生的呢？通过非致死性突变分析模型的建立与应用, 产生了新的适应性进化观点, 即环境胁迫诱导细胞适应性突变。这种环境诱导的细胞突变过程涉及多方面的生理调控, 包括细胞内毒性物质(如氧活性物质)积累并造成DNA损伤、DNA错配修复的活性受到抑制、胞内RpoS反应和SOS反应被激活等。这些反应使胞内高保真的DNA复制状态转变为低保真的DNA修复状态, 提高胞内突变率和重组活性。此外, 基因转录影响基因组的不稳定, 容易产生DNA损伤, 并造成局部的高突变率, 即形成了转录偶联的DNA修复与突变为基础的适应性突变观点。文章围绕环境胁迫诱导细胞突变率增加和转录偶联的DNA修复与突变这两种适应性突变分子机制, 阐述其相关的研究进展, 以期更好地理解环境条件诱导细胞发生适应性突变的过程。     

Ecologically relevant stress resistance: from microarrays and quantitative trait loci to candidate genes — A research plan and preliminary results using<Emphasis Type="Italic">Drosophila</Emphasis> as a model organism and climatic and genetic stress as model stresses

We aim at studying adaptation to genetic and environmental stress and its evolutionary implications at different levels of biological organization. Stress influences cellular processes, individual physiology, genetic variation at the population level, and the process of natural selection. To investigate these highly connected levels of stress effects, it is advisable - if not critical - to integrate approaches from ecology, evolution, physiology, molecular biology and genetics. To investigate the mechanisms of stress resistance, how resistance evolves, and what factors contribute to and constrain its evolution, we use the well-defined model systems ofDrosophila species, representing both cosmopolitan species such asD. melanogaster with a known genome map, and more specialized and ecologically well described species such as the cactophilicD. buzzatii. Various climate-related stresses are used as model stresses including desiccation, starvation, cold and heat. Genetic stress or genetic load is modelled by studying the consequences of inbreeding, the accumulation of (slightly) deleterious mutations, hybridization or the loss of genetic variability. We present here a research plan and preliminary results combining various approaches: molecular techniques such as microarrays, quantitative trait loci (QTL) analyses, quantitative PCR, ELISA or Western blotting are combined with population studies of resistance to climatic and genetic stress in natural populations collected across climatic gradients as well as in selection lines maintained in the laboratory.     

Par force evolution as a mechanism for rapid adaptation

An important task is to reduce the cost of natural selection. The stress provides a short-term non-specific resistance to a number of factors (cross resistance); however, in the long term, it is harmful (distress). In a small population, it is more expedient to change the stress flow (prolonging the phase of cross resistance, neutralizing distress) by natural selection within a small group of stress genes than to test all mutations of genome in a search for preadaptations. This process can provoke allo- or aromorphosis.     

Whole Genome,Whole Population Sequencing Reveals That Loss of Signaling Networks Is the Major Adaptive Strategy in a Constant Environment

Molecular signaling networks are ubiquitous across life and likely evolved to allow organisms to sense and respond to environmental change in dynamic environments. Few examples exist regarding the dispensability of signaling networks, and it remains unclear whether they are an essential feature of a highly adapted biological system. Here, we show that signaling network function carries a fitness cost in yeast evolving in a constant environment. We performed whole-genome, whole-population Illumina sequencing on replicate evolution experiments and find the major theme of adaptive evolution in a constant environment is the disruption of signaling networks responsible for regulating the response to environmental perturbations. Over half of all identified mutations occurred in three major signaling networks that regulate growth control: glucose signaling, Ras/cAMP/PKA and HOG. This results in a loss of environmental sensitivity that is reproducible across experiments. However, adaptive clones show reduced viability under starvation conditions, demonstrating an evolutionary tradeoff. These mutations are beneficial in an environment with a constant and predictable nutrient supply, likely because they result in constitutive growth, but reduce fitness in an environment where nutrient supply is not constant. Our results are a clear example of the myopic nature of evolution: a loss of environmental sensitivity in a constant environment is adaptive in the short term, but maladaptive should the environment change.     

Bacterial Adaptation through Loss of Function

The metabolic capabilities and regulatory networks of bacteria have been optimized by evolution in response to selective pressures present in each species'' native ecological niche. In a new environment, however, the same bacteria may grow poorly due to regulatory constraints or biochemical deficiencies. Adaptation to such conditions can proceed through the acquisition of new cellular functionality due to gain of function mutations or via modulation of cellular networks. Using selection experiments on transposon-mutagenized libraries of bacteria, we illustrate that even under conditions of extreme nutrient limitation, substantial adaptation can be achieved solely through loss of function mutations, which rewire the metabolism of the cell without gain of enzymatic or sensory function. A systematic analysis of similar experiments under more than 100 conditions reveals that adaptive loss of function mutations exist for many environmental challenges. Drawing on a wealth of examples from published articles, we detail the range of mechanisms through which loss-of-function mutations can generate such beneficial regulatory changes, without the need for rare, specific mutations to fine-tune enzymatic activities or network connections. The high rate at which loss-of-function mutations occur suggests that null mutations play an underappreciated role in the early stages of adaption of bacterial populations to new environments.     

Clonal Interference in the Evolution of Influenza

The seasonal influenza A virus undergoes rapid evolution to escape human immune response. Adaptive changes occur primarily in antigenic epitopes, the antibody-binding domains of the viral hemagglutinin. This process involves recurrent selective sweeps, in which clusters of simultaneous nucleotide fixations in the hemagglutinin coding sequence are observed about every 4 years. Here, we show that influenza A (H3N2) evolves by strong clonal interference. This mode of evolution is a red queen race between viral strains with different beneficial mutations. Clonal interference explains and quantifies the observed sweep pattern: we find an average of at least one strongly beneficial amino acid substitution per year, and a given selective sweep has three to four driving mutations on average. The inference of selection and clonal interference is based on frequency time series of single-nucleotide polymorphisms, which are obtained from a sample of influenza genome sequences over 39 years. Our results imply that mode and speed of influenza evolution are governed not only by positive selection within, but also by background selection outside antigenic epitopes: immune adaptation and conservation of other viral functions interfere with each other. Hence, adapting viral proteins are predicted to be particularly brittle. We conclude that a quantitative understanding of influenza’s evolutionary and epidemiological dynamics must be based on all genomic domains and functions coupled by clonal interference.