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.     

Relating fitness to long‐term environmental variations in natura

Quantifying links between ecological processes and adaptation dynamics in natura remains a crucial challenge. Many studies have documented the strength, form and direction of selection, and its variations in space and time, but only a few managed to link these variations to their proximal causes. This step is, however, crucial, if we are to understand how the variation in selective pressure affects adaptive allele dynamics in natural settings. We used data from a long‐term survey (about 30 years) monitoring the adaptation to insecticides of Culex pipiens mosquitoes in Montpellier area (France), focusing on three resistance alleles of the Ester locus. We used a population genetics model taking temporal and spatial variations in selective pressure into account, to assess the quantitative relationships between variations in the proximal agent of selection (amounts of insecticide sprayed) and the fitness of resistance alleles. The response to variations in selective pressure was fast, and the alleles displayed different fitness‐to‐environment relationships: the analyses revealed that even slight changes in insecticide doses could induce changes in the strength and direction of selection, thus changing the fitness ranking of the adaptive alleles. They also revealed that selective pressures other than the insecticides used for mosquito control affected the resistance allele dynamics. These fitness‐to‐environment relationships, fast responses and continuous evolution limit our ability to predict the outcome of adaptive allele dynamics in a changing environment, but they clearly contribute to the maintenance of polymorphism in natural populations. Our study also emphasizes the necessity of long‐term surveys in evolutionary ecology.     

Loci under selection during multiple range expansions of an invasive plant are mostly population specific,but patterns are associated with climate

Identifying the genes underlying rapid evolutionary changes, describing their function and ascertaining the environmental pressures that determine fitness are the central elements needed for understanding of evolutionary processes and phenotypic changes that improve the fitness of populations. It has been hypothesized that rapid adaptive changes in new environments may contribute to the rapid spread and success of invasive plants and animals. As yet, studies of adaptation during invasion are scarce, as is knowledge of the genes underlying adaptation, especially in multiple replicated invasions. Here, we quantified how genotype frequencies change during invasions, resulting in rapid evolution of naturalized populations. We used six fully replicated common garden experiments in Brazil where Pinus taeda (loblolly pine) was introduced at the same time, in the same numbers, from the same seed sources, and has formed naturalized populations expanding outward from the plantations. We used a combination of nonparametric, population genetics and multivariate statistics to detect changes in genotype frequencies along each of the six naturalization gradients and their association with climate as well as shifts in allele frequencies compared to the source populations. Results show 25 genes with significant shifts in genotype frequencies. Six genes had shifts in more than one population. Climate explained 25% of the variation in the groups of genes under selection across all locations, but specific genes under strong selection during invasions did not show climate‐related convergence. In conclusion, we detected rapid evolutionary changes during invasive range expansions, but the particular gene‐level patterns of evolution may be population specific.     

Multigenerational versus single generation studies to estimate herbicide resistance fitness cost in Arabidopsis thaliana

The evolution of resistance in response to pesticide selection is expected to be delayed if fitness costs are associated with resistance genes. The estimate of fitness costs usually involves comparing major growth traits of resistant versus susceptible individuals in the absence of pesticide. Ideally, a measure of changes in resistance allele frequency over several generations would allow the best estimate of the overall fitness cost of a resistance gene. In greenhouse conditions, we monitored the dynamics of the evolution of the frequencies of six herbicide-resistant mutations (acetolactate synthase, cellulose synthase, and auxin-induced target genes) in the model species Arabidopsis thaliana in a multigenerational study covering five to seven nonoverlapping generations. The microevolutionary dynamics in experimental populations indicated a mean fitness cost of 38%, 73%, and 94% for the ixr1-2, axr1-3, and axr2-1 resistances, respectively; no fitness cost for the csr1-1, and ixr2-1 resistances; and a transient advantage for the aux1-7 resistance. The result for the csr1-1 resistance contrasts with a cost of 37% based on total seed number in a previous study, demonstrating that single generation studies could have limitation for detecting cost. A positive frequency dependence for the fitness cost was also detected for the ixr1-2 resistance. The results are discussed in relation to the maintenance of polymorphism at resistance loci.     

Insecticide resistance in the mosquito Culex pipiens: what have we learned about adaptation?

Resistance to organophosphate (OP) insecticide in the mosquito Culex pipiens has been studied for ca. 30 years. This example of micro-evolution has been thoroughly investigated as an opportunity to assess precisely both the new adapted phenotypes and the associated genetic changes. A notable feature is that OP resistance is achieved with few genes, and these genes have generally large effects. The molecular events generating such resistance genes are complex (e.g., gene amplification, gene regulation) potentially explaining their low frequency of de novo occurrence. In contrast, migration is a frequent event, including passive transportation between distant populations. This generates a complex interaction between mutations and migration, and promotes competition among resistance alleles. When the precise physiological action of each gene product is rather well known, it is possible to understand the dominance level or the type of epistasis observed. It is however difficult to predict a priori how resistance genes will interact, and it is too early to state whether or not this will be ever possible. These resistance genes are costly, and the cost is variable among them. It is usually believed that the initial fitness cost would gradually decrease due to subsequent mutations with a modifier effect. In the present example, a particular modifier occurred (a gene duplication) at one resistance locus, whereas at the other one reduction of cost is driven by allele replacement and apparently not by selection of modifiers.     

Parallel trait adaptation across opposing thermal environments in experimental Drosophila melanogaster populations

Thermal stress is a pervasive selective agent in natural populations that impacts organismal growth, survival, and reproduction. Drosophila melanogaster exhibits a variety of putatively adaptive phenotypic responses to thermal stress in natural and experimental settings; however, accompanying assessments of fitness are typically lacking. Here, we quantify changes in fitness and known thermal tolerance traits in replicated experimental D. melanogaster populations following more than 40 generations of evolution to either cyclic cold or hot temperatures. By evaluating fitness for both evolved populations alongside a reconstituted starting population, we show that the evolved populations were the best adapted within their respective thermal environments. More strikingly, the evolved populations exhibited increased fitness in both environments and improved resistance to both acute heat and cold stress. This unexpected parallel response appeared to be an adaptation to the rapid temperature changes that drove the cycling thermal regimes, as parallel fitness changes were not observed when tested in a constant thermal environment. Our results add to a small, but growing group of studies that demonstrate the importance of fluctuating temperature changes for thermal adaptation and highlight the need for additional work in this area.     

Resistance Gene Replacement in the Mosquito Culex pipiens: Fitness Estimation From Long-Term Cline Series

How adaptation appears and is later refined by natural selection has been the object of intense theoretical work. However, the testing of these theories is limited by our ability to estimate the strength of natural selection in nature. Using a long-term cline series, we estimate the selection coefficients acting on different alleles at the same locus to analyze the allele replacement observed in the insecticide resistance gene Ester in the mosquito Culex pipiens in the Montpellier area, southern France. Our method allows us to accurately account for the resistance allele replacement observed in this area since 1986. A first resistance allele appeared early, which was replaced by a second resistance allele providing the same advantage but at a lower cost, itself being replaced by a third resistance allele with both higher advantage and cost. It shows that amelioration of the adaptation (here resistance to insecticide) through allele replacement was successively achieved by selection of first a generalist allele (i.e., with a low fitness variance across environments) and later a specialist allele (i.e., with a large fitness variance across environments). More generally, we discuss how precise estimates of the strength of selection obtained from field data help us understand the process of amelioration of adaptation.     

Temperature‐sensitive fitness cost of insecticide resistance in Chinese populations of the diamondback moth Plutella xylostella

Alleles conferring a higher adaptive value in one environment may have a detrimental impact on fitness in another environment. Alleles conferring resistance to pesticides and drugs provide textbook examples of this trade‐off as, in addition to conferring resistance to these molecules, they frequently decrease fitness in pesticide/drug‐free environments. We show here that resistance to chlorpyrifos, an organophosphate (OP), in Chinese populations of the diamondback moth, Plutella xylostella, is conferred by two mutations of ace1 – the gene encoding the acetylcholinesterase enzyme targeted by OPs – affecting the amino acid sequence of the corresponding protein. These mutations were always linked, consistent with the segregation of a single resistance allele, ace1R, carrying both mutations, in the populations studied. We monitored the frequency of ace1R (by genotyping more than 20 000 adults) and the level of resistance (through bioassays on more than 50 000 individuals) over several generations. We found that the ace1R resistance allele was costly in the absence of insecticide and that this cost was likely recessive. This fitness costs involved a decrease in fecundity: females from resistant strains laid 20% fewer eggs, on average, than females from susceptible strains. Finally, we found that the fitness costs associated with the ace1R allele were greater at high temperatures. At least two life history traits were involved: longevity and fecundity. The relative longevity of resistant individuals was affected only at high temperatures and the relative fecundity of resistant females – which was already affected at temperatures optimal for development – decreased further at high temperatures. The implications of these findings for resistance management are discussed.     

Escaping an evolutionary lobster trap: drug resistance and compensatory mutation in a fluctuating environment

The fitness landscape concept aids intuition on adaptive evolution through low fitness genotypes. Evolutionary processes become complex when environments and therefore fitnesses fluctuate. Antibiotic resistance evolution in bacteria is an important example of such dynamics. Resistance bears a cost in the drug-free environment, but compensatory mutation can lower this cost, creating a fitness valley. With the drug present, the valley becomes a hill that is easily climbed. Once a population is dominated by resistant-compensated genotypes, reversion to sensitivity is difficult: this phenomenon has been described as an evolutionary lobster trap. With increasing frequencies of drug resistance among pathogenic bacteria, it is critical to understand how this trap can be escaped. Here, we develop stochastic models to investigate these dynamics. The residual fitness cost (the cost remaining after compensatory mutation has occurred) is a key parameter. Reversion to sensitivity is favored when the time spent in the absence of the drug relative to its presence is high compared to the residual fitness cost. Population sizes are also important: in large populations, resistant-compensated mutants appear in resistant-uncompensated or sensitive-compensated genotypes without fixation of these intermediates. This stochastic tunneling effect occurs when sufficient time is allowed by the rates of environmental fluctuation.     

Constraints on adaptive mutations in the codling moth Cydia pomonella (L.): measuring fitness trade-offs and natural selection

Adaptive changes in populations encountering a new environment are often constrained by deleterious pleiotropic interactions with ancestral physiological functions. Evolutionary responses of populations can thus be limited by natural selection under fluctuating environmental conditions, if the adaptive mutations are associated with pleiotropic fitness costs. In this context, we have followed the evolution of the frequencies of insecticide-resistant mutants of Cydia pomonella when reintroduced into an untreated environment. The novel set of selective forces after removal of insecticide pressure led to the decline of the frequencies of resistant phenotypes over time, suggesting that the insecticide-adapted genetic variants were selected against the absence of insecticide (with a selective coefficient estimated at 0.11). The selective coefficients were also estimated for both the major cytochrome P450-dependent monooxygenase (MFO) and the minor glutathione S-transferase (GST) systems (0.17 and negligible, respectively), which have been previously shown to be involved in resistance. The involvement of metabolic systems acting both through xenobiotic detoxification and biosynthetic pathways of endogenous compounds may be central to explaining the deleterious physiological consequences resulting from pleiotropy of adaptive changes. The estimation of the magnitude of the fitness cost associated with insecticide resistance in C. pomonella suggests that resistance management strategies exclusively based on insecticide alternations would be unlikely to delay such a selection process.     

Stressful environments can indirectly select for increased longevity

Longevity is modulated by a range of conserved genes in eukaryotes, but it is unclear how variation in these genes contributes to the evolution of longevity in nature. Mutations that increase life span in model organisms typically induce trade‐offs which lead to a net reduction in fitness, suggesting that such mutations are unlikely to become established in natural populations. However, the fitness consequences of manipulating longevity have rarely been assessed in heterogeneous environments, in which stressful conditions are encountered. Using laboratory selection experiments, we demonstrate that long‐lived, stress‐resistant Caenorhabditis elegans age‐1(hx546) mutants have higher fitness than the wild‐type genotype if mixed genotype populations are periodically exposed to high temperatures when food is not limited. We further establish, using stochastic population projection models, that the age‐1(hx546) mutant allele can confer a selective advantage if temperature stress is encountered when food availability also varies over time. Our results indicate that heterogeneity in environmental stress may lead to altered allele frequencies over ecological timescales and indirectly drive the evolution of longevity. This has important implications for understanding the evolution of life‐history strategies.     

Recurrent adaptation in a low‐dispersal trait

The study of natural populations from contrasting environments has greatly enhanced our understanding of ecological‐dependent selection, adaptation and speciation. Cases of parallel evolution in particular have facilitated the study of the molecular and genetic basis of adaptive variation. This includes the type and number of genes underlying adaptive traits, as well as the extent to which these genes are exchanged among populations and contribute repeatedly to parallel evolution. Yet, surprisingly few studies provide a comprehensive view on the evolutionary history of adaptive traits from mutation to widespread adaptation. When did key mutations arise, how did they increase in frequency, and how did they spread? In this issue of Molecular Ecology, Van Belleghem et al. ( 2015 ) reconstruct the evolutionary history of a gene associated with wing size in the salt marsh beetle Pogonus chalceus. Screening the entire distribution range of this species, they found a single origin for the allele associated with the short‐winged ecotype. This allele seemingly evolved in an isolated population and rapidly introgressed into other populations. These findings suggest that the adaptive genetic variation found in sympatric short‐ and long‐winged populations has an allopatric origin, confirming that allopatric phases may be important at early stages of speciation.     

Rapid genome‐wide evolution in Brassica rapa populations following drought revealed by sequencing of ancestral and descendant gene pools

There is increasing evidence that evolution can occur rapidly in response to selection. Recent advances in sequencing suggest the possibility of documenting genetic changes as they occur in populations, thus uncovering the genetic basis of evolution, particularly if samples are available from both before and after selection. Here, we had a unique opportunity to directly assess genetic changes in natural populations following an evolutionary response to a fluctuation in climate. We analysed genome‐wide differences between ancestors and descendants of natural populations of Brassica rapa plants from two locations that rapidly evolved changes in multiple phenotypic traits, including flowering time, following a multiyear late‐season drought in California. These ancestor‐descendant comparisons revealed evolutionary shifts in allele frequencies in many genes. Some genes showing evolutionary shifts have functions related to drought stress and flowering time, consistent with an adaptive response to selection. Loci differentiated between ancestors and descendants (F_ST outliers) were generally different from those showing signatures of selection based on site frequency spectrum analysis (Tajima's D), indicating that the loci that evolved in response to the recent drought and those under historical selection were generally distinct. Very few genes showed similar evolutionary responses between two geographically distinct populations, suggesting independent genetic trajectories of evolution yielding parallel phenotypic changes. The results show that selection can result in rapid genome‐wide evolutionary shifts in allele frequencies in natural populations, and highlight the usefulness of combining resurrection experiments in natural populations with genomics for studying the genetic basis of adaptive evolution.     

The Population and Evolutionary Dynamics of Homologous Gene Recombination in Bacteria

In bacteria, recombination is a rare event, not a part of the reproductive process. Nevertheless, recombination—broadly defined to include the acquisition of genes from external sources, i.e., horizontal gene transfer (HGT)—plays a central role as a source of variation for adaptive evolution in many species of bacteria. Much of niche expansion, resistance to antibiotics and other environmental stresses, virulence, and other characteristics that make bacteria interesting and problematic, is achieved through the expression of genes and genetic elements obtained from other populations of bacteria of the same and different species, as well as from eukaryotes and archaea. While recombination of homologous genes among members of the same species has played a central role in the development of the genetics and molecular biology of bacteria, the contribution of homologous gene recombination (HGR) to bacterial evolution is not at all clear. Also, not so clear are the selective pressures responsible for the evolution and maintenance of transformation, the only bacteria-encoded form of HGR. Using a semi-stochastic simulation of mutation, recombination, and selection within bacterial populations and competition between populations, we explore (1) the contribution of HGR to the rate of adaptive evolution in these populations and (2) the conditions under which HGR will provide a bacterial population a selective advantage over non-recombining or more slowly recombining populations. The results of our simulation indicate that, under broad conditions: (1) HGR occurring at rates in the range anticipated for bacteria like Streptococcus pneumoniae, Escherichia coli, Haemophilus influenzae, and Bacillus subtilis will accelerate the rate at which a population adapts to environmental conditions; (2) once established in a population, selection for this capacity to increase rates of adaptive evolution can maintain bacteria-encoded mechanisms of recombination and prevent invasion of non-recombining populations, even when recombination engenders a modest fitness cost; and (3) because of the density- and frequency-dependent nature of HGR in bacteria, this capacity to increase rates of adaptive evolution is not sufficient as a selective force to provide a recombining population a selective advantage when it is rare. Under realistic conditions, homologous gene recombination will increase the rate of adaptive evolution in bacterial populations and, once established, selection for higher rates of evolution will promote the maintenance of bacteria-encoded mechanisms for HGR. On the other hand, increasing rates of adaptive evolution by HGR is unlikely to be the sole or even a dominant selective pressure responsible for the original evolution of transformation.     

Environmental variation alters the fitness effects of rifampicin resistance mutations in Pseudomonas aeruginosa

The fitness effects of antibiotic resistance mutations in antibiotic‐free conditions play a key role in determining the long‐term maintenance of resistance. Although resistance is usually associated with a cost, the impact of environmental variation on the cost of resistance is poorly understood. Here, we test the impact of heterogeneity in temperature and resource availability on the fitness effects of antibiotic resistance using strains of the pathogenic bacterium Pseudomonas aeruginosa carrying clinically important rifampicin resistance mutations. Although the rank order of fitness was generally maintained across environments, fitness effects relative to the wild type differed significantly. Changes in temperature had a profound impact on the fitness effects of resistance, whereas changes in carbon substrate had only a weak impact. This suggests that environmental heterogeneity may influence whether the costs of resistance are likely to be ameliorated by second‐site compensatory mutations or by reversion to wild‐type rpoB. Our results highlight the need to consider environmental heterogeneity and genotype‐by‐environment interactions for fitness in models of resistance evolution.     

Fitness cost due to herbicide resistance may trigger genetic background evolution

This article investigates the possible existence of mechanisms counterbalancing the negative pleiotropic effects on development and reproduction that are conferred by alleles responsible for herbicide resistance in the weed Alopecurus myosuroides. We considered three herbicide-resistant, mutant acetyl-coenzyme A carboxylase (ACCase) alleles, Leu1781, Asn2041, and Gly2078, found in eight resistant populations. Of these, Gly2078 is the only allele with a known fitness cost. We compared plants homozygous for wild-type ACCase alleles that were siblings of plants carrying a given mutant resistant ACCase allele with plants from three populations where resistance did not evolve. In each of two series of experiments, we measured germination dynamics, seedling vigor, plant height, vegetative biomass, and seed production. The wild-type siblings of plants carrying Gly2078 performed better in the field, on average, than wild-type plants that were sibling of plants carrying other mutant ACCase alleles, and particularly those carrying Leu1781. We propose that rapid evolution of the genetic background of plants from the populations where the Gly2078 allele originally arose could partially counterbalance Gly2078 fitness cost, enhancing the spread of the resistant genotypes.     

Detecting genetic responses to environmental change

Changes in environmental conditions can rapidly shift allele frequencies in populations of species with relatively short generation times. Frequency shifts might be detectable in neutral genetic markers when stressful conditions cause a population decline. However, frequency shifts that are diagnostic of specific conditions depend on isolating sets of genes that are involved in adaptive responses. Shifts at candidate loci underlying adaptive responses and DNA regions that control their expression have now been linked to evolutionary responses to pollution, global warming and other changes. Conversely, adaptive constraints, particularly in physiological traits, are recognized through DNA decay in candidate genes. These approaches help researchers and conservation managers understand the power and constraints of evolution.     

Insecticide resistance genes induce a mating competition cost in Culex pipiens mosquitoes

Resistance to organophosphorus insecticides (OP) in Culex pipiens mosquitoes represents a convenient model for investigating the fitness cost of resistance genes and its origin, since both the environmental changes in nature and the adaptive genes are clearly identified. Two loci are involved in this resistance--the super-locus Ester and the locus Ace.1--each displaying several resistance alleles. Population surveys have shown differences in fitness cost between these resistance genes and even between resistance alleles of the same locus. In order to better understand this fitness cost and its variability, the effects of these resistance genes on several fitness-related traits are being studied. Here, through competition experiments between two males for the access to one female, we analysed the effect on paternity success associated with three resistance alleles--Ester4, Ester1 and Ace.1R--relative to susceptible males and relative to one another. The eventual effect of female genotype on male mating success was also studied by using susceptible and resistant females. The strains used in this experiment had the same genetic background. Susceptible males had a mating advantage when competing with any of the resistant males, suggesting a substantial cost of resistance genes to this trait. When competing against susceptible males, the paternity success did not vary among resistant males, whatever the genotype of the female. When competing against other resistant males, no difference in paternity success was apparent, except when the female was Ester1.     

Parasite‐mediated selection in a natural metapopulation of Daphnia magna

Parasite‐mediated selection varying across time and space in metapopulations is expected to result in host local adaptation and the maintenance of genetic diversity in disease‐related traits. However, nonadaptive processes like migration and extinction‐(re)colonization dynamics might interfere with adaptive evolution. Understanding how adaptive and nonadaptive processes interact to shape genetic variability in life‐history and disease‐related traits can provide important insights into their evolution in subdivided populations. Here we investigate signatures of spatially fluctuating, parasite‐mediated selection in a natural metapopulation of Daphnia magna. Host genotypes from infected and uninfected populations were genotyped at microsatellite markers, and phenotyped for life‐history and disease traits in common garden experiments. Combining phenotypic and genotypic data a Q_ST–F_ST‐like analysis was conducted to test for signatures of parasite mediated selection. We observed high variation within and among populations for phenotypic traits, but neither an indication of host local adaptation nor a cost of resistance. Infected populations have a higher gene diversity (Hs) than uninfected populations and Hs is strongly positively correlated with fitness. These results suggest a strong parasite effect on reducing population level inbreeding. We discuss how stochastic processes related to frequent extinction‐(re)colonization dynamics as well as host and parasite migration impede the evolution of resistance in the infected populations. We suggest that the genetic and phenotypic patterns of variation are a product of dynamic changes in the host gene pool caused by the interaction of colonization bottlenecks, inbreeding, immigration, hybrid vigor, rare host genotype advantage and parasitism. Our study highlights the effect of the parasite in ameliorating the negative fitness consequences caused by the high drift load in this metapopulation.