Threespine Stickleback: A Model System For Evolutionary Genomics.

The repeated adaptation of oceanic threespine sticklebacks to fresh water has made it a premier organism to study parallel evolution. These small fish have multiple distinct ecotypes that display a wide range of diverse phenotypic traits. Ecotypes are easily crossed in the laboratory, and families are large and develop quickly enough for quantitative trait locus analyses, positioning the threespine stickleback as a versatile model organism to address a wide range of biological questions. Extensive genomic resources, including linkage maps, a high-quality reference genome, and developmental genetics tools have led to insights into the genomic basis of adaptation and the identification of genomic changes controlling traits in vertebrates. Recently, threespine sticklebacks have been used as a model system to identify the genomic basis of highly complex traits, such as behavior and host-microbiome and host-parasite interactions. We review the latest findings and new avenues of research that have led the threespine stickleback to be considered a supermodel of evolutionary genomics.

Genomic landscapes of divergence among island bird populations: Evidence of parallel adaptation but at different loci?

When populations colonise new environments, they may be exposed to novel selection pressures but also suffer from extensive genetic drift due to founder effects, small population sizes and limited interpopulation gene flow. Genomic approaches enable us to study how these factors drive divergence, and disentangle neutral effects from differentiation at specific loci due to selection. Here, we investigate patterns of genetic diversity and divergence using whole-genome resequencing (>22× coverage) in Berthelot's pipit (Anthus berthelotii), a passerine endemic to the islands of three north Atlantic archipelagos. Strong environmental gradients, including in pathogen pressure, across populations in the species range, make it an excellent system in which to explore traits important in adaptation and/or incipient speciation. First, we quantify how genomic divergence accumulates across the speciation continuum, that is, among Berthelot's pipit populations, between sub species across archipelagos, and between Berthelot's pipit and its mainland ancestor, the tawny pipit (Anthus campestris). Across these colonisation timeframes (2.1 million-ca. 8000 years ago), we identify highly differentiated loci within genomic islands of divergence and conclude that the observed distributions align with expectations for non-neutral divergence. Characteristic signatures of selection are identified in loci associated with craniofacial/bone and eye development, metabolism and immune response between population comparisons. Interestingly, we find limited evidence for repeated divergence of the same loci across the colonisation range but do identify different loci putatively associated with the same biological traits in different populations, likely due to parallel adaptation. Incipient speciation across these island populations, in which founder effects and selective pressures are strong, may therefore be repeatedly associated with morphology, metabolism and immune defence.

The phoenix hypothesis of speciation.

Genetic divergence among allopatric populations builds reproductive isolation over time. This process is accelerated when populations face a changing environment that allows large-effect mutational differences to accumulate, but abrupt change also places populations at risk of extinction. Here we use simulations of Fisher's geometric model with explicit population dynamics to explore the genetic changes that occur in the face of environmental changes. Because evolutionary rescue leads to the fixation of mutations whose phenotypic effects are larger on average compared with populations not at risk of extinction, these mutations are thus more likely to lead to reproductive isolation. We refer to the formation of new species from the ashes of populations in decline as the phoenix hypothesis of speciation. The phoenix hypothesis predicts more substantial hybrid fitness breakdown among populations surviving a higher extinction risk. The hypothesis was supported when many loci underlie adaptation. With only a small number of potential rescue mutations, however, mutations that fixed in different populations were more likely to be identical, with such parallel changes reducing isolation. Consequently, reproductive isolation builds fastest in populations subject to an intermediate extinction risk, given a limited number of mutations available for adaptation.

Mutation bias can shape adaptation in large asexual populations experiencing clonal interference.

The extended evolutionary synthesis invokes a role for development in shaping adaptive evolution, which in population genetics terms corresponds to mutation-biased adaptation. Critics have claimed that clonal interference makes mutation-biased adaptation rare. We consider the behaviour of two simultaneously adapting traits, one with larger mutation rate U, the other with larger selection coefficient s, using asexual travelling wave models. We find that adaptation is dominated by whichever trait has the faster rate of adaptation v in isolation, with the other trait subject to evolutionary stalling. Reviewing empirical claims for mutation-biased adaptation, we find that not all occur in the 'origin-fixation' regime of population genetics where v is only twice as sensitive to s as to U. In some cases, differences in U are at least ten to twelve times larger than differences in s, as needed to cause mutation-biased adaptation even in the 'multiple mutations' regime. Surprisingly, when U > s in the 'diffusive-mutation' regime, the required sensitivity ratio is also only two, despite pervasive clonal interference. Given two traits with identical v, the benefit of having higher s is surprisingly small, occurring largely when one trait is at the boundary between the origin-fixation and multiple mutations regimes.

Independent domestication events in the blue-cheese fungus Penicillium roqueforti.

Domestication provides an excellent framework for studying adaptive divergence. Using population genomics and phenotypic assays, we reconstructed the domestication history of the blue cheese mould Penicillium roqueforti. We showed that this fungus was domesticated twice independently. The population used in Roquefort originated from an old domestication event associated with weak bottlenecks and exhibited traits beneficial for pre-industrial cheese production (slower growth in cheese and greater spore production on bread, the traditional multiplication medium). The other cheese population originated more recently from the selection of a single clonal lineage, was associated with all types of blue cheese worldwide except Roquefort, and displayed phenotypes more suited for industrial cheese production (high lipolytic activity, efficient cheese cavity colonization ability and salt tolerance). We detected genomic regions affected by recent positive selection and putative horizontal gene transfers. This study sheds light on the processes of rapid adaptation and raises questions about genetic resource conservation.

Non-parallel divergence across freshwater and marine three-spined stickleback Gasterosteus aculeatus populations.

This work investigated whether multiple freshwater populations of three-spined stickleback Gasterosteus aculeatus in different freshwater catchments in the Jutland Peninsula, Denmark, derived from the same marine populations show repeated adaptive responses. A total of 327 G. aculeatus collected at 13 sampling locations were screened for genetic variation using a combination of 70 genes putatively under selection and 26 neutral genes along with a marker linked to the ectodysplasin gene (eda), which is strongly correlated with plate armour morphs in the species. A highly significant genetic differentiation was found that was higher among different freshwater samples than between marine-freshwater samples. Tests for selection between marine and freshwater populations showed a very low degree of parallelism and no single nucleotide polymorphism was detected as outlier in all freshwater-marine pairwise comparisons, including the eda. This suggests that G. aculeatus is not necessarily the prime example of parallel local adaptation suggested in much of the literature and that important exceptions exist (i.e. the Jutland Peninsula). While marine populations in the results described here showed a high phenotype-genotype correlation at eda, a low association was found for most of the freshwater populations. The most extreme case was found in the freshwater Lake Hald where all low-plated phenotypes were either homozygotes for the allele supposed to be associated with completely plated morphs or heterozygotes, but none were homozygotes for the putative low-plated allele. Re-examination of data from seven G. aculeatus studies agrees in showing a high but partial association between phenotype-genotype at eda in G. aculeatus freshwater populations and that mismatches occur everywhere in the European regions studied (higher in some areas, i.e. Denmark). This is independent of the eda marker used.

Evidence for extensive parallelism but divergent genomic architecture of adaptation along altitudinal and latitudinal gradients in Populus trichocarpa.

Adaptation to climate across latitude and altitude reflects shared climatic constraints, which may lead to parallel adaptation. However, theory predicts that higher gene flow should favor more concentrated genomic architectures, which would lead to fewer locally maladapted recombinants. We used exome capture to resequence the gene space along a latitudinal and two altitudinal transects in the model tree Populus trichocapra. Adaptive trait phenotyping was coupled with FST outlier tests and sliding window analysis to assess the degree of parallel adaptation as well as the genomic distribution of outlier loci. Up to 51% of outlier loci overlapped between transect pairs and up to 15% of these loci overlapped among all three transects. Genomic clustering of adaptive loci was more pronounced for altitudinal than latitudinal transects. In both altitudinal transects, there was a larger number of these 'islands of divergence', which were on average longer and included several of exceptional physical length. Our results suggest that recapitulation of genetic clines over latitude and altitude involves extensive parallelism, but that steep altitudinal clines generate islands of divergence. This suggests that physical proximity of genes in coadapted complexes may buffer against the movement of maladapted alleles from geographically proximal but climatically distinct populations.

Detecting parallel polygenic adaptation to novel evolutionary pressure in wild populations: a case study in Atlantic cod (Gadus morhua).

Populations can adapt to novel selection pressures through dramatic frequency changes in a few genes of large effect or subtle shifts in many genes of small effect. The latter (polygenic adaptation) is expected to be the primary mode of evolution for many life-history traits but tends to be more difficult to detect than changes in genes of large effect. Atlantic cod (Gadus morhua) were subjected to intense fishing pressure over the twentieth century, leading to abundance crashes and a phenotypic shift toward earlier maturation across many populations. Here, we use spatially replicated temporal genomic data to test for a shared polygenic adaptive response to fishing using methods previously applied to evolve-and-resequence experiments. Cod populations on either side of the Atlantic show covariance in allele frequency change across the genome that are characteristic of recent polygenic adaptation. Using simulations, we demonstrate that the degree of covariance in allele frequency change observed in cod is unlikely to be explained by neutral processes or background selection. As human pressures on wild populations continue to increase, understanding and attributing modes of adaptation using methods similar to those demonstrated here will be important in identifying the capacity for adaptive responses and evolutionary rescue. This article is part of the theme issue 'Detecting and attributing the causes of biodiversity change: needs, gaps and solutions'.

Domestication in dry-cured meat Penicillium fungi: Convergent specific phenotypes and horizontal gene transfers without strong genetic subdivision.

Some fungi have been domesticated for food production, with genetic differentiation between populations from food and wild environments, and food populations often acquiring beneficial traits through horizontal gene transfers (HGTs). Studying their adaptation to human-made substrates is of fundamental and applied importance for understanding adaptation processes and for further strain improvement. We studied here the population structures and phenotypes of two distantly related Penicillium species used for dry-cured meat production, P. nalgiovense, the most common species in the dry-cured meat food industry, and P. salamii, used locally by farms. Both species displayed low genetic diversity, lacking differentiation between strains isolated from dry-cured meat and those from other environments. Nevertheless, the strains collected from dry-cured meat within each species displayed slower proteolysis and lipolysis than their wild conspecifics, and those of P. nalgiovense were whiter. Phenotypically, the non-dry-cured meat strains were more similar to their sister species than to their conspecific dry-cured meat strains, indicating an evolution of specific phenotypes in dry-cured meat strains. A comparison of available Penicillium genomes from various environments revealed HGTs, particularly between P. nalgiovense and P. salamii (representing almost 1.5 Mb of cumulative length). HGTs additionally involved P. biforme, also found in dry-cured meat products. We further detected positive selection based on amino acid changes. Our findings suggest that selection by humans has shaped the P. salamii and P. nalgiovense populations used for dry-cured meat production, which constitutes domestication. Several genetic and phenotypic changes were similar in P. salamii, P. nalgiovense and P. biforme, indicating convergent adaptation to the same human-made environment. Our findings have implications for fundamental knowledge on adaptation and for the food industry: the discovery of different phenotypes and of two mating types paves the way for strain improvement by conventional breeding, to elucidate the genomic bases of beneficial phenotypes and to generate diversity.

Parallel and population-specific gene regulatory evolution in cold-adapted fly populations.

Changes in gene regulation at multiple levels may comprise an important share of the molecular changes underlying adaptive evolution in nature. However, few studies have assayed within- and between-population variation in gene regulatory traits at a transcriptomic scale, and therefore inferences about the characteristics of adaptive regulatory changes have been elusive. Here, we assess quantitative trait differentiation in gene expression levels and alternative splicing (intron usage) between three closely related pairs of natural populations of Drosophila melanogaster from contrasting thermal environments that reflect three separate instances of cold tolerance evolution. The cold-adapted populations were known to show population genetic evidence for parallel evolution at the SNP level, and here we find evidence for parallel expression evolution between them, with stronger parallelism at larval and adult stages than for pupae. We also implement a flexible method to estimate cis- vs trans-encoded contributions to expression or splicing differences at the adult stage. The apparent contributions of cis- vs trans-regulation to adaptive evolution vary substantially among population pairs. While two of three population pairs show a greater enrichment of cis-regulatory differences among adaptation candidates, trans-regulatory differences are more likely to be implicated in parallel expression changes between population pairs. Genes with significant cis-effects are enriched for signals of elevated genetic differentiation between cold- and warm-adapted populations, suggesting that they are potential targets of local adaptation. These findings expand our knowledge of adaptive gene regulatory evolution and our ability to make inferences about this important and widespread process.

Population genomics supports clonal reproduction and multiple independent gains and losses of parasitic abilities in the most devastating nematode pest.

The root-knot nematodes are the most devastating worms to worldwide agriculture with Meloidogyne incognita being the most widely distributed and damaging species. This parasitic and ecological success seems surprising given its supposed obligatory clonal reproduction. Clonal reproduction has been suspected based on cytological observations but, so far, never confirmed by population genomics data. As a species, M. incognita is highly polyphagous with thousands of host plants. However, different M. incognita isolates present distinct and overlapping patterns of host compatibilities. Historically, four "host races" had been defined as a function of ranges of compatible and incompatible plants. In this study, we used population genomics to assess whether (a) reproduction is actually clonal in this species, (b) the host races follow an underlying phylogenetic signal or, rather represent multiple independent transitions, and (c) how genome variations associate with other important biological traits such as the affected crops and geographical distribution. We sequenced the genomes of 11 M. incognita isolates across Brazil that covered the four host races in replicates. By aligning the genomic reads of these isolates to the M. incognita reference genome assembly, we identified point variations. Analysis of linkage disequilibrium and 4-gametes test showed no evidence for recombination, corroborating the clonal reproduction of M. incognita. The few point variations between the isolates showed no significant association with the host races, the geographical origin of the samples, or the crop on which they have been collected. Addition of isolates from other locations around the world confirmed this lack of underlying phylogenetic signal. This suggests multiple gains and losses of parasitic abilities and adaptations to different environments account for the broad host spectrum and wide geographical distribution of M. incognita and thus to its high economic impact. This surprising adaptability without sex poses both evolutionary and agro-economic challenges.

Adaptive Horizontal Gene Transfers between Multiple Cheese-Associated Fungi.

Domestication is an excellent model for studies of adaptation because it involves recent and strong selection on a few, identified traits [1-5]. Few studies have focused on the domestication of fungi, with notable exceptions [6-11], despite their importance to bioindustry [12] and to a general understanding of adaptation in eukaryotes [5]. Penicillium fungi are ubiquitous molds among which two distantly related species have been independently selected for cheese making-P. roqueforti for blue cheeses like Roquefort and P. camemberti for soft cheeses like Camembert. The selected traits include morphology, aromatic profile, lipolytic and proteolytic activities, and ability to grow at low temperatures, in a matrix containing bacterial and fungal competitors [13-15]. By comparing the genomes of ten Penicillium species, we show that adaptation to cheese was associated with multiple recent horizontal transfers of large genomic regions carrying crucial metabolic genes. We identified seven horizontally transferred regions (HTRs) spanning more than 10 kb each, flanked by specific transposable elements, and displaying nearly 100% identity between distant Penicillium species. Two HTRs carried genes with functions involved in the utilization of cheese nutrients or competition and were found nearly identical in multiple strains and species of cheese-associated Penicillium fungi, indicating recent selective sweeps; they were experimentally associated with faster growth and greater competitiveness on cheese and contained genes highly expressed in the early stage of cheese maturation. These findings have industrial and food safety implications and improve our understanding of the processes of adaptation to rapid environmental changes.

Too much of a good thing: the unique and repeated paths toward copper adaptation.

Copper is a micronutrient essential for growth due to its role as a cofactor in enzymes involved in respiration, defense against oxidative damage, and iron uptake. Yet too much of a good thing can be lethal, and yeast cells typically do not have tolerance to copper levels much beyond the concentration in their ancestral environment. Here, we report a short-term evolutionary study of Saccharomyces cerevisiae exposed to levels of copper sulfate that are inhibitory to the initial strain. We isolated and identified adaptive mutations soon after they arose, reducing the number of neutral mutations, to determine the first genetic steps that yeast take when adapting to copper. We analyzed 34 such strains through whole-genome sequencing and by assaying fitness within different environments; we also isolated a subset of mutations through tetrad analysis of four lines. We identified a multilayered evolutionary response. In total, 57 single base-pair mutations were identified across the 34 lines. In addition, gene amplification of the copper metallothionein protein, CUP1-1, was rampant, as was chromosomal aneuploidy. Four other genes received multiple, independent mutations in different lines (the vacuolar transporter genes VTC1 and VTC4; the plasma membrane H+-ATPase PMA1; and MAM3, a protein required for normal mitochondrial morphology). Analyses indicated that mutations in all four genes, as well as CUP1-1 copy number, contributed significantly to explaining variation in copper tolerance. Our study thus finds that evolution takes both common and less trodden pathways toward evolving tolerance to an essential, but highly toxic, micronutrient.

Longitudinal trends in climate drive flowering time clines in North American Arabidopsis thaliana.

Introduced species frequently show geographic differentiation, and when differentiation mirrors the ancestral range, it is often taken as evidence of adaptive evolution. The mouse-ear cress (Arabidopsis thaliana) was introduced to North America from Eurasia 150-200 years ago, providing an opportunity to study parallel adaptation in a genetic model organism. Here, we test for clinal variation in flowering time using 199 North American (NA) accessions of A. thaliana, and evaluate the contributions of major flowering time genes FRI, FLC, and PHYC as well as potential ecological mechanisms underlying differentiation. We find evidence for substantial within population genetic variation in quantitative traits and flowering time, and putatively adaptive longitudinal differentiation, despite low levels of variation at FRI, FLC, and PHYC and genome-wide reductions in population structure relative to Eurasian (EA) samples. The observed longitudinal cline in flowering time in North America is parallel to an EA cline, robust to the effects of population structure, and associated with geographic variation in winter precipitation and temperature. We detected major effects of FRI on quantitative traits associated with reproductive fitness, although the haplotype associated with higher fitness remains rare in North America. Collectively, our results suggest the evolution of parallel flowering time clines through novel genetic mechanisms.