Replicate Populations Research Articles

Shared genetic architecture links energy metabolism, behavior and starvation resistance along a power-endurance axis

Abstract Shared developmental, physiological, and molecular mechanisms can generate strong genetic covariances across suites of traits, constraining genetic variability, and evolvability to certain axes in multivariate trait space (“variational modules” or “syndromes”). Such trait suites will not only respond jointly to selection; they will also covary across populations that diverged from one another by genetic drift. We report evidence for such a genetically correlated trait suite that links traits related to energy metabolism along a “power-endurance” axis in Drosophila melanogaster. The “power” pole of the axis is characterized by high potential for energy generation and expenditure—high expression of glycolysis and TCA cycle genes, high abundance of mitochondria, and high spontaneous locomotor activity. The opposite “endurance” pole is characterized by high triglyceride (fat) reserves, locomotor endurance, and starvation resistance (and low values of traits associated with the “power” pole). This trait suite also aligns with the first principal component of metabolome; the “power” direction is characterized by low levels of trehalose (blood sugar) and high levels of some amino acids and their derivatives, including creatine, a compound known to facilitate energy production in muscles. Our evidence comes from six replicate “Selected” populations adapted to a nutrient-poor larval diet regime during 250 generations of experimental evolution and six “Control” populations evolved in parallel on a standard diet regime. We found that, within each of these experimental evolutionary regimes, the above traits strongly covaried along this “power-endurance” axis across replicate populations which diversified by drift, indicating a shared genetic architecture. The two evolutionary regimes also drove divergence along this axis, with Selected populations on average displaced towards the “power” direction compared to Controls. Aspects of this “power-endurance” axis resemble the “pace of life” syndrome and the “thrifty phenotype”; it may have evolved as part of a coordinated organismal response to nutritional conditions.

Temporal dynamics of scent mark composition in field‐experimental lizard populations

Abstract Animal signals are dynamic traits that can undergo considerable spatial and temporal changes and that are influenced by factors such as age, health condition and interactions with both the abiotic and biotic environment. However, much of our understanding of signal changes throughout an individual's lifetime stems from cross‐sectional, often laboratory‐based, studies focused on visual and auditory signals. Longitudinal field investigations of temporal variation in chemical signals, especially in vertebrates, remain rare despite chemical communication being the most ubiquitous form of information exchange in the natural world. To remedy this, we conducted a unique, replicated field experiment to study the temporal signal dynamics in free‐living lizard populations on natural islands. Specifically, we collected scent marks from individually marked lizards across five populations during the spring of two consecutive years and analysed the lipophilic chemical composition of these scent marks. Our findings demonstrate that the overall scent mark composition of individual lizards changed over time, shifting consistently in both direction and magnitude from year to year among individuals and across replicate populations. Similar patterns were observed for the chemical richness and diversity of scent marks. Temporal variation in the relative proportions of three potentially socially relevant signalling compounds in lizard scent marks revealed a more complex pattern: α‐tocopherol remained stable over time, oleic acid decreased and the change in octadecanoic acid proportion was body size‐dependent. Together, our results provide novel insights into how individual vertebrate chemical signals may fluctuate across space and time. We discuss the potential causes of the observed temporal variability and its consequences for chemical signal evolution. Read the free Plain Language Summary for this article on the Journal blog.

Evolution in microbial microcosms is highly parallel, regardless of the presence of interacting species

Evolution often follows similar trajectories in replicate populations, suggesting that it may be predictable. However, populations are naturally embedded in multispecies communities, and the extent to which evolution is contingent on the specific species interacting with the focal population is still largely unexplored. Here, we study adaptations in strains of 11 different species, experimentally evolved both in isolation and in various pairwise co-cultures. Although partner-specific effects are detectable, evolution was mostly shared between strains evolved with different partners; similar changes occurred in strains' growth abilities, in community properties, and in about half of the repeatedly mutated genes. This pattern persisted even in species pre-adapted to the abiotic conditions. These findings indicate that evolution may not always depend strongly on the biotic environment, making predictions regarding coevolutionary dynamics less challenging than previously thought. A record of this paper's transparent peer review process is included in the supplemental information.

Histone H3-K27 and H3-K9 methylation does not predict lifespan in divergent populations of Drosophila melanogaster

A universal theory of aging remains elusive despite decades of research. Traditional gerontological research has focused on genetic alterations as the driving unit of the aging processes. Recent breakthroughs in cellular reprogramming, transgenerational epigenetic inheritance, and molecular aging clocks have motivated theories proposing that the disruption of epigenetic regulation is the proximal cause of aging. The claim that aging is fundamentally an epigenetic process, however, is largely based on studies using clonal cell populations and genetically identical organisms. Consequently, there has been limited experimental evidence demonstrating the correlation between epigenetic differences and longevity differences among genetically diverse populations. To address this gap in literature we leveraged laboratory populations of the model organism Drosophila melanogaster. Using sister populations with evolved differences in longevity after more than 1 500 generations of divergent life history selection, we measured the methylation at the H3K27 and H3K9 sites which have previously been shown to become dysregulated with age. Here we report that there were no differences in the amount of H3K27me3 or H3K9me2 between 3 replicate population pairs of the divergent lines. These results do not support that epigenetic dysregulation is the underlying cause of aging. Future studies investigating additional indicators of epigenetic dysregulation are required to clarify the role of epigenetics in aging.

Evolution recovers the fitness of Acinetobacter baylyi strains with large deletions through mutations in deletion-specific targets and global post-transcriptional regulators.

Organelles and endosymbionts have naturally evolved dramatically reduced genome sizes compared to their free-living ancestors. Synthetic biologists have purposefully engineered streamlined microbial genomes to create more efficient cellular chassis and define the minimal components of cellular life. During natural or engineered genome streamlining, deletion of many non-essential genes in combination often reduces bacterial fitness for idiosyncratic or unknown reasons. We investigated how and to what extent laboratory evolution could overcome these defects in six variants of the transposon-free Acinetobacter baylyi strain ADP1-ISx that each had a deletion of a different 22- to 42-kilobase region and two strains with larger deletions of 70 and 293 kilobases. We evolved replicate populations of ADP1-ISx and each deletion strain for ~300 generations in a chemically defined minimal medium or a complex medium and sequenced the genomes of endpoint clonal isolates. Fitness increased in all cases that were examined except for two ancestors that each failed to improve in one of the two environments. Mutations affecting nine protein-coding genes and two small RNAs were significantly associated with one of the two environments or with certain deletion ancestors. The global post-transcriptional regulators rnd (ribonuclease D), csrA (RNA-binding carbon storage regulator), and hfq (RNA-binding protein and chaperone) were frequently mutated across all strains, though the incidence and effects of these mutations on gene function and bacterial fitness varied with the ancestral deletion and evolution environment. Mutations in this regulatory network likely compensate for how an earlier deletion of a transposon in the ADP1-ISx ancestor of all the deletion strains restored csrA function. More generally, our results demonstrate that fitness lost during genome streamlining can usually be regained rapidly through laboratory evolution and that recovery tends to occur through a combination of deletion-specific compensation and global regulatory adjustments.

Host and antibiotic jointly select for greater virulence in Staphylococcus aureus.

Widespread antibiotic usage has resulted in the rapid evolution of drug-resistant bacterial pathogens and poses significant threats to public health. Resolving how pathogens respond to antibiotics under different contexts is critical for understanding disease emergence and evolution going forward. The impact of antibiotics has been demonstrated most directly through in vitro pathogen passaging experiments. Independent from antibiotic selection, interactions with hosts have also altered the evolutionary trajectories and fitness landscapes of pathogens, shaping infectious disease outcomes. However, it is unclear how interactions between hosts and antibiotics impact the evolution of pathogen virulence. Here, we evolved and re-sequenced Staphylococcus aureus , a major bacterial pathogen, varying exposure to host and antibiotics to tease apart the contributions of these selective pressures on pathogen adaptation. After 12 passages, S. aureus evolving in Caenorhabditis elegans nematodes exposed to a sub-minimum inhibitory concentration of antibiotic (oxacillin) became highly virulent, regardless of whether the ancestral pathogen was methicillin-resistant (MRSA) or methicillin-sensitive (MSSA). Host and antibiotic exposure selected for reduced drug susceptibility in MSSA lineages while increasing MRSA total growth outside hosts. We identified mutations in genes involved in complex regulatory networks linking virulence and metabolism, including codY , agr , and gdpP , suggesting that rapid adaptation to infect hosts may have pleiotropic effects. In particular, MSSA populations under selection from host and antibiotic accumulated mutations in the global regulator gene codY , which controls biofilm formation in S. aureus. These populations had indeed evolved more robust biofilms-a trait linked to both virulence and antibiotic resistance-suggesting evolution of one trait can confer multiple adaptive benefits. Despite evolving in similar environments, MRSA and MSSA populations proceeded on divergent evolutionary paths, with MSSA populations exhibiting more similarities across replicate populations. Our results underscore the importance of considering multiple and concurrent selective pressures as drivers of pervasive pathogen traits.

Experimental evolution suggests rapid assembly of the 'selfing syndrome' from standing variation in Mimulus guttatus.

Ecological and evolutionary changes are likely to occur rapidly when outcrossing populations experience pollinator loss. However, the number and identify of plant traits that will respond to this form of selection, as well as the overall predictability of evolutionary responses, remain unclear. We experimentally evolved 20 large replicate populations of Mimulus guttatus for 10 generations under three treatments: pure outcrossing, mixed mating (10% outcrossing) and pure selfing. These populations were founded from the same genetically diverse and outcrossing natural population. After 10 generations, all measured traits evolved with flower size, phenology, and reproductive traits diverging consistently among mating system treatments. Autogamy increased dramatically in the selfing treatment, but the magnitude of adaptation only becomes clear once inbreeding depression is factored out. Selfing treatment plants evolved reduced stigma-anther separation, and also exhibited declines in flower size and per-flower reproductive capacity. Flower size also declined in selfing populations but this was driven mainly by inbreeding depression and cannot be attributed to adaptation towards the selfing syndrome. Generally, the mixed mating populations evolved trait values intermediate to the fully selfing and outcrossing populations. Overall, our experimental treatments reiterated differences that have been documented in interspecific comparisons between selfing and outcrossing species pairs. Given that such contrasts involve species separated by thousands or even millions of generations, it is noteworthy that large evolutionary responses were obtained from genetic variation segregating within a single natural population.

Mutations in yeast are deleterious on average regardless of the degree of adaptation to the testing environment.

The role of spontaneous mutations in evolution depends on the distribution of their effects on fitness. Despite a general consensus that new mutations are deleterious on average, a handful of mutation accumulation experiments in diverse organisms instead suggest that beneficial and deleterious mutations can have comparable fitness impacts, i.e. the product of their respective rates and effects can be roughly equal. We currently lack a general framework for predicting when such a pattern will occur. One idea is that beneficial mutations will be more evident in genotypes that are not well adapted to the testing environment. We tested this prediction experimentally in the laboratory yeast Saccharomyces cerevisiae by allowing nine replicate populations to adapt to novel environments with complex sets of stressors. After >1000 asexual generations interspersed with 41 rounds of sexual reproduction, we assessed the mean effect of induced mutations on yeast growth in both the environment to which they had been adapting and the alternative novel environment. The mutations were deleterious on average, with the severity depending on the testing environment. However, we found no evidence that the adaptive match between genotype and environment is predictive of mutational fitness effects.

Reproductive isolation arises during laboratory adaptation to a novel hot environment

BackgroundReproductive isolation can result from adaptive processes (e.g., ecological speciation and mutation-order speciation) or stochastic processes such as “system drift” model. Ecological speciation predicts barriers to gene flow between populations from different environments, but not among replicate populations from the same environment. In contrast, reproductive isolation among populations independently adapted to the same/similar environment can arise from both mutation-order speciation or system drift.ResultsIn experimentally evolved populations adapting to a hot environment for over 100 generations, we find evidence for pre- and postmating reproductive isolation. On one hand, an altered lipid metabolism and cuticular hydrocarbon composition pointed to possible premating barriers between the ancestral and replicate evolved populations. On the other hand, the pronounced gene expression differences in male reproductive genes may underlie the postmating isolation among replicate evolved populations adapting to the same environment with the same standing genetic variation.ConclusionOur study confirms that replicated evolution experiments provide valuable insights into the mechanisms of speciation. The rapid emergence of the premating reproductive isolation during temperature adaptation showcases incipient ecological speciation. The potential evidence of postmating reproductive isolation among replicates gave rise to two hypotheses: (1) mutation-order speciation through a common selection on early fecundity leading to an inherent inter-locus sexual conflict; (2) system drift with genetic drift along the neutral ridges.

Stochastic parabolic growth promotes coexistence and a relaxed error threshold in RNA-like replicator populations

The RNA world hypothesis proposes that during the early evolution of life, primordial genomes of the first self-propagating evolutionary units existed in the form of RNA-like polymers. Autonomous, non-enzymatic, and sustained replication of such information carriers presents a problem, because product formation and hybridization between template and copy strands reduces replication speed. Kinetics of growth is then parabolic with the benefit of entailing competitive coexistence, thereby maintaining diversity. Here, we test the information-maintaining ability of parabolic growth in stochastic multispecies population models under the constraints of constant total population size and chemostat conditions. We find that large population sizes and small differences in the replication rates favor the stable coexistence of the vast majority of replicator species (‘genes’), while the error threshold problem is alleviated relative to exponential amplification. In addition, sequence properties (GC content) and the strength of resource competition mediated by the rate of resource inflow determine the number of coexisting variants, suggesting that fluctuations in building block availability favored repeated cycles of exploration and exploitation. Stochastic parabolic growth could thus have played a pivotal role in preserving viable sequences generated by random abiotic synthesis and providing diverse genetic raw material to the early evolution of functional ribozymes.

Stochastic parabolic growth promotes coexistence and a relaxed error threshold in RNA-like replicator populations.

The RNA world hypothesis proposes that during the early evolution of life, primordial genomes of the first self-propagating evolutionary units existed in the form of RNA-like polymers. Autonomous, non-enzymatic, and sustained replication of such information carriers presents a problem, because product formation and hybridization between template and copy strands reduces replication speed. Kinetics of growth is then parabolic with the benefit of entailing competitive coexistence, thereby maintaining diversity. Here, we test the information-maintaining ability of parabolic growth in stochastic multispecies population models under the constraints of constant total population size and chemostat conditions. We find that large population sizes and small differences in the replication rates favor the stable coexistence of the vast majority of replicator species ('genes'), while the error threshold problem is alleviated relative to exponential amplification. In addition, sequence properties (GC content) and the strength of resource competition mediated by the rate of resource inflow determine the number of coexisting variants, suggesting that fluctuations in building block availability favored repeated cycles of exploration and exploitation. Stochastic parabolic growth could thus have played a pivotal role in preserving viable sequences generated by random abiotic synthesis and providing diverse genetic raw material to the early evolution of functional ribozymes.

A Robust Growth-Based Selection Platform to Evolve an Enzyme via Dependency on Noncanonical Tyrosine Analogues.

Growth-based selections evaluate the fitness of individual organisms at a population level. In enzyme engineering, such growth selections allow for the rapid and straightforward identification of highly efficient biocatalysts from extensive libraries. However, selection-based improvement of (synthetically useful) biocatalysts is challenging, as they require highly dependable strategies that artificially link their activities to host survival. Here, we showcase a robust and scalable growth-based selection platform centered around the complementation of noncanonical amino acid-dependent bacteria. Specifically, we demonstrate how serial passaging of populations featuring millions of carbamoylase variants autonomously selects biocatalysts with up to 90,000-fold higher initial rates. Notably, selection of replicate populations enriched diverse biocatalysts, which feature distinct amino acid motifs that drastically boost carbamoylase activity. As beneficial substitutions also originated from unintended copying errors during library preparation or cell division, we anticipate that our growth-based selection platform will be applicable to the continuous, autonomous evolution of diverse biocatalysts in the future.

Polymorphisms in transcription factor binding sites and enhancer regions and pancreatic ductal adenocarcinoma risk

Genome-wide association studies (GWAS) are a powerful tool for detecting variants associated with complex traits and can help risk stratification and prevention strategies against pancreatic ductal adenocarcinoma (PDAC). However, the strict significance threshold commonly used makes it likely that many true risk loci are missed. Functional annotation of GWAS polymorphisms is a proven strategy to identify additional risk loci. We aimed to investigate single-nucleotide polymorphisms (SNP) in regulatory regions [transcription factor binding sites (TFBSs) and enhancers] that could change the expression profile of multiple genes they act upon and thereby modify PDAC risk. We analyzed a total of 12,636 PDAC cases and 43,443 controls from PanScan/PanC4 and the East Asian GWAS (discovery populations), and the PANDoRA consortium (replication population). We identified four associations that reached study-wide statistical significance in the overall meta-analysis: rs2472632(A) (enhancer variant, OR 1.10, 95%CI 1.06,1.13, p = 5.5 × 10−8), rs17358295(G) (enhancer variant, OR 1.16, 95%CI 1.10,1.22, p = 6.1 × 10−7), rs2232079(T) (TFBS variant, OR 0.88, 95%CI 0.83,0.93, p = 6.4 × 10−6) and rs10025845(A) (TFBS variant, OR 1.88, 95%CI 1.50,1.12, p = 1.32 × 10−5). The SNP with the most significant association, rs2472632, is located in an enhancer predicted to target the coiled-coil domain containing 34 oncogene. Our results provide new insights into genetic risk factors for PDAC by a focused analysis of polymorphisms in regulatory regions and demonstrating the usefulness of functional prioritization to identify loci associated with PDAC risk.

Experimental evolution of hybrid populations to identify Dobzhansky-Muller incompatibility loci.

Epistatic interactions between loci that reduce fitness in interspecies hybrids, Dobzhansky-Muller incompatibilities (DMIs), contribute genetically to the inviability and infertility within hybrid populations. It remains a challenge, however, to identify the loci that contribute to DMIs as causes of reproductive isolation between species. Here, we assess through forward simulation the power of evolve-and-resequence (E&R) experimental evolution of hybrid populations to map DMI loci. We document conditions under which such a mapping strategy may be most feasible and demonstrate how mapping power is sensitive to biologically relevant parameters such as one-way versus two-way incompatibility type, selection strength, recombination rate, and dominance interactions. We also assess the influence of parameters under direct control of an experimenter, including duration of experimental evolution and number of replicate populations. We conclude that an E&R strategy for mapping DMI loci, and other cases of epistasis, can be a viable option under some circumstances for study systems with short generation times like Caenorhabditis nematodes.

Phenotypic stasis with genetic divergence.

Whether or not genetic divergence in the short-term of tens to hundreds of generations is compatible with phenotypic stasis remains a relatively unexplored problem. We evolved predominantly outcrossing, genetically diverse populations of the nematode Caenorhabditis elegans under a constant and homogeneous environment for 240 generations and followed individual locomotion behavior. Although founders of lab populations show highly diverse locomotion behavior, during lab evolution, the component traits of locomotion behavior - defined as the transition rates in activity and direction - did not show divergence from the ancestral population. In contrast, transition rates' genetic (co)variance structure showed a marked divergence from the ancestral state and differentiation among replicate populations during the final 100 generations and after most adaptation had been achieved. We observe that genetic differentiation is a transient pattern during the loss of genetic variance along phenotypic dimensions under drift during the last 100 generations of lab evolution. These results suggest that short-term stasis of locomotion behavior is maintained because of stabilizing selection, while the genetic structuring of component traits is contingent upon drift history.

The impact of interspecific competition on the genomic evolution of Phaeobacter inhibens and Pseudoalteromonas tunicata during biofilm growth.

Interspecific interactions in biofilms have been shown to cause the emergence of community-level properties. To understand the impact of interspecific competition on evolution, we deep-sequenced the dispersal population of mono- and co-culture biofilms of two antagonistic marine bacteria (Phaeobacter inhibens 2.10 and Pseudoalteromononas tunicata D2). Enhanced phenotypic and genomic diversification was observed in the P. tunicata D2 populations under both mono- and co-culture biofilms in comparison to P. inhibens 2.10. The genetic variation was exclusively due to single nucleotide variants and small deletions, and showed high variability between replicates, indicating their random emergence. Interspecific competition exerted an apparent strong positive selection on a subset of P. inhibens 2.10 genes (e.g., luxR, cobC, argH, and sinR) that could facilitate competition, while the P. tunicata D2 population was genetically constrained under competition conditions. In the absence of interspecific competition, the P. tunicata D2 replicate populations displayed high levels of mutations affecting the same genes involved in cell motility and biofilm formation. Our results show that interspecific biofilm competition has a complex impact on genomic diversification, which likely depends on the nature of the competing strains and their ability to generate genetic variants due to their genomic constraints.

Adaptive DNA amplification of synthetic gene circuit opens a way to overcome cancer chemoresistance.

Drug resistance continues to impede the success of cancer treatments, creating a need for experimental model systems that are broad, yet simple, to allow the identification of mechanisms and novel countermeasures applicable to many cancer types. To address these needs, we investigated a set of engineered mammalian cell lines with synthetic gene circuits integrated into their genome that evolved resistance to Puromycin. We identified DNA amplification as the mechanism underlying drug resistance in 4 out of 6 replicate populations. Triplex-forming oligonucleotide (TFO) treatment combined with Puromycin could efficiently suppress the growth of cell populations with DNA amplification. Similar observations in human cancer cell lines suggest that TFOs could be broadly applicable to mitigate drug resistance, one of the major difficulties in treating cancer.

Ploidy evolution in a wild yeast is linked to an interaction between cell type and metabolism

Ploidy is an evolutionarily labile trait, and its variation across the tree of life has profound impacts on evolutionary trajectories and life histories. The immediate consequences and molecular causes of ploidy variation on organismal fitness are frequently less clear, although extreme mating type skews in some fungi hint at links between cell type and adaptive traits. Here, we report an unusual recurrent ploidy reduction in replicate populations of the budding yeast Saccharomyces eubayanus experimentally evolved for improvement of a key metabolic trait, the ability to use maltose as a carbon source. We find that haploids have a substantial, but conditional, fitness advantage in the absence of other genetic variation. Using engineered genotypes that decouple the effects of ploidy and cell type, we show that increased fitness is primarily due to the distinct transcriptional program deployed by haploid-like cell types, with a significant but smaller contribution from absolute ploidy. The link between cell-type specification and the carbon metabolism adaptation can be traced to the noncanonical regulation of a maltose transporter by a haploid-specific gene. This study provides novel mechanistic insight into the molecular basis of an environment–cell type fitness interaction and illustrates how selection on traits unexpectedly linked to ploidy states or cell types can drive karyotypic evolution in fungi.

Spatial replication can best advance our understanding of population responses to climate

Understanding the responses of plant populations dynamics to climatic variability is frustrated by the need for long‐term datasets. Here, we advocate for new studies that estimate the effects of climate by sampling replicate populations in locations with similar climate. We first use data analysis on spatial locations in the conterminous USA to assess how far apart spatial replicates should be from each other to minimize temporal correlations in climate. We find that on average spatial locations separated by 316 km (SD = 149 km) have moderate (0.5) correlations in annual precipitation. Second, we use simulations to demonstrate that spatial replication can lead to substantial gains in the range of climates sampled during a given set of years so long as the climate correlations between the populations are at low to moderate levels. Third, we use simulations to quantify how many spatial replicates would be necessary to achieve the same statistical power of a single‐population, long‐term data set under different strengths and directions of spatial correlations in climate between spatial replicates. Our results indicate that spatial replication is an untapped opportunity to study the effects of climate on demography and to rapidly fill important knowledge gaps in the field of population ecology.

Selection on plastic adherence leads to hyper-multicellular strains and incidental virulence in the budding yeast.

Many disease-causing microbes are not obligate pathogens; rather, they are environmental microbes taking advantage of an ecological opportunity. The existence of microbes whose life cycle does not require a host and are not normally pathogenic, yet are well-suited to host exploitation, is an evolutionary puzzle. One hypothesis posits that selection in the environment may favor traits that incidentally lead to pathogenicity and virulence, or serve as pre-adaptations for survival in a host. An example of such a trait is surface adherence. To experimentally test the idea of 'accidental virulence', replicate populations of Saccharomyces cerevisiae were evolved to attach to a plastic bead for hundreds of generations. Along with plastic adherence, two multicellular phenotypes- biofilm formation and flor formation- increased; another phenotype, pseudohyphal growth, responded to the nutrient limitation. Thus, experimental selection led to the evolution of highly-adherent, hyper-multicellular strains. Wax moth larvae injected with evolved hyper-multicellular strains were significantly more likely to die than those injected with evolved non-multicellular strains. Hence, selection on plastic adherence incidentally led to the evolution of enhanced multicellularity and increased virulence. Our results support the idea that selection for a trait beneficial in the open environment can inadvertently generate opportunistic, 'accidental' pathogens.