Instances of parallel ecotypic divergence where adaptation to similar conditions repeatedly cause similar phenotypic changes in closely related organisms are useful for studying the role of ecological selection in speciation. Here we used a combination of traditional and next generation genotyping techniques to test for the parallel divergence of plants from the Senecio lautus complex, a phenotypically variable groundsel that has adapted to disparate environments in the South Pacific. Phylogenetic analysis of a broad selection of Senecio species showed that members of the S. lautus complex form a distinct lineage that has diversified recently in Australasia. An inspection of thousands of polymorphisms in the genome of 27 natural populations from the S. lautus complex in Australia revealed a signal of strong genetic structure independent of habitat and phenotype. Additionally, genetic differentiation between populations was correlated with the geographical distance separating them, and the genetic diversity of populations strongly depended on geographical location. Importantly, coastal forms appeared in several independent phylogenetic clades, a pattern that is consistent with the parallel evolution of these forms. Analyses of the patterns of genomic differentiation between populations further revealed that adjacent populations displayed greater genomic heterogeneity than allopatric populations and are differentiated according to variation in soil composition. These results are consistent with a process of parallel ecotypic divergence in face of gene flow.

Adaptation to replicate environments is often achieved through similar phenotypic solutions. Whether selection also produces convergent genomic changes in these situations remains largely unknown. The variable groundsel, Senecio lautus, is an excellent system to investigate the genetic underpinnings of convergent evolution, because morphologically similar forms of these plants have adapted to the same environments along the coast of Australia. We compared range-wide patterns of genomic divergence in natural populations of this plant and searched for regions putatively affected by natural selection. Our results indicate that environmental adaptation followed complex genetic trajectories, affecting multiple loci, implying both the parallel recruitment of the same alleles and the divergence of completely different genomic regions across geography. An analysis of the biological functions of candidate genes suggests that adaptation to coastal environments may have occurred through the recruitment of different genes participating in similar processes. The relatively low genetic convergence that characterizes the parallel evolution of S. lautus forms suggests that evolution is more constrained at higher levels of biological organization.

Summary Speciation with gene flow, or the evolution of reproductive isolation between interbreeding populations, remains a controversial problem in evolution. This is because gene flow erodes the adaptive differences that selection creates between populations. Here, we use a combination of common garden experiments in the field and in the glasshouse to investigate what ecological and genetic mechanisms prevent gene flow and maintain morphological and genetic differentiation between coastal parapatric populations of the Australian groundsel Senecio lautus. We discovered that in each habitat extrinsic reproductive barriers prevented gene flow, whereas intrinsic barriers in F1 hybrids were weak. In the field, herbivores played a major role in preventing gene flow, but glasshouse experiments demonstrated that soil type also created variable selective pressures both locally and on a greater geographic scale. Our experimental results demonstrate that interfertile plant populations adapting to contrasting environments may diverge as a consequence of concurrent natural selection acting against migrants and hybrids through multiple mechanisms. These results provide novel insights into the consequences of local adaptation in the origin of strong barriers to gene flow in plants, and suggest that herbivory may play an important role in the early stages of plant speciation.

Adaptation to contrasting environments across a heterogeneous landscape favors the formation of ecotypes by promoting ecological divergence. Patterns of fitness variation in the field can show whether natural selection drives local adaptation and ecotype formation. However, to demonstrate a link between ecological divergence and speciation, local adaptation must have consequences for reproductive isolation. Using contrasting ecotypes of an Australian wildflower, Senecio lautus in common garden experiments, hybridization experiments, and reciprocal transplants, we assessed how the environment shapes patterns of adaptation and the consequences of adaptive divergence for reproductive isolation. Local adaptation was strong between ecotypes, but weaker between populations of the same ecotype. F1 hybrids exhibited heterosis, but crosses involving one native parent performed better than those with two foreign parents. In a common garden experiment, F2 hybrids exhibited reduced fitness compared to parentals and F1 hybrids, suggesting that few genetic incompatibilities have accumulated between populations adapted to contrasting environments. Our results show how ecological differences across the landscape have created complex patterns of local adaptation and reproductive isolation, suggesting that divergent natural selection has played a fundamental role in the early stages of species diversification.

Genetic correlations between traits can bias adaptation away from optimal phenotypes and constrain the rate of evolution. If genetic correlations between traits limit adaptation to contrasting environments, rapid adaptive divergence across a heterogeneous landscape may be difficult. However, if genetic variance can evolve and align with the direction of natural selection, then abundant allelic variation can promote rapid divergence during adaptive radiation. Here, we explored adaptive divergence among ecotypes of an Australian native wildflower by quantifying divergence in multivariate phenotypes of populations that occupy four contrasting environments. We investigated differences in multivariate genetic variance underlying morphological traits and examined the alignment between divergence in phenotype and divergence in genetic variance. We found that divergence in mean multivariate phenotype has occurred along two major axes represented by different combinations of plant architecture and leaf traits. Ecotypes also showed divergence in the level of genetic variance in individual traits, and the multivariate distribution of genetic variance among traits. Divergence in multivariate phenotypic mean aligned with divergence in genetic variance, with most of the divergence in phenotype among ecotypes associated with a change in trait combinations that had substantial levels of genetic variance in each ecotype. Overall, our results suggest that divergent natural selection acting on high levels of standing genetic variation might fuel ecotypic differentiation during the early stages of adaptive radiation.peer-reviewed)

Genetic correlations between traits can concentrate genetic variance into fewer phenotypic dimensions that can bias evolutionary trajectories along the axis of greatest genetic variance and away from optimal phenotypes, constraining the rate of evolution. If genetic correlations limit adaptation, rapid adaptive divergence between multiple contrasting environments may be difficult. However, if natural selection increases the frequency of rare alleles after colonization of new environments, an increase in genetic variance in the direction of selection can accelerate adaptive divergence. Here, we explored adaptive divergence of an Australian native wildflower by examining the alignment between divergence in phenotype mean and divergence in genetic variance among four contrasting ecotypes. We found divergence in mean multivariate phenotype along two major axes represented by different combinations of plant architecture and leaf traits. Ecotypes also showed divergence in the level of genetic variance in individual traits and the multivariate distribution of genetic variance among traits. Divergence in multivariate phenotypic mean aligned with divergence in genetic variance, with much of the divergence in phenotype among ecotypes associated with changes in trait combinations containing substantial levels of genetic variance. Overall, our results suggest that natural selection can alter the distribution of genetic variance underlying phenotypic traits, increasing the amount of genetic variance in the direction of natural selection and potentially facilitating rapid adaptive divergence during an adaptive radiation.

19Natural selection is a major driver for the origins of adaptations and new species 1 . Whether or 20 not the processes driving adaptation and speciation share a molecular basis remains largely 21 unknown 2 . Here, we show that divergence in hormone signalling contributed to the evolution 22 habits of S. lautus populations, therefore making evolution of the auxin pathway a natural 60 candidate to link the molecular basis of adaptation and speciation. We reasoned that if 61 divergence in the auxin pathway contributed to the evolution of adaptation and speciation in 62 S. lautus, we would discover the following evidence: First, we would detect similar patterns 63 of genetic divergence in auxin-related pathways across multiple erect and prostrate hybrid 64 and natural populations. Second, these populations would differ in phenotypes dependent on 65 auxin, such as their ability to alter the direction of growth in relation to gravity 10,16 . And third, 66 divergence in these auxin-dependent phenotypes would contribute to local adaptation and 67 intrinsic reproductive isolation between populations. 68We test these hypotheses primarily on coastal populations of S. lautus (Fig. 1a, Extended 69Data Table 1), which exhibit strong correlations between growth habit and the environments 70 they occupy 7 . Populations inhabiting sand dunes (Dune hereafter) are erect, while populations 71 growing on adjacent rocky headlands (Headland hereafter) are prostrate ( Fig. 1b). Erect and 72 prostrate growth habits can also be found in related populations from the alpine regions of 73 Australia, with a prostrate population inhabiting an exposed alpine meadow and an erect 74 population inhabiting a sheltered alpine gully (Fig. 1c). Dune populations are continually 75 exposed to high temperatures and sun radiation, low salinity, and low nutrient sand substrate, 76whereas Headland populations are exposed to high salinity, high nutrients and powerful 77 winds 17 . Neighbouring Dune and Headland populations are often sister taxa, group into two 78 major monophyletic clades (eastern and south-eastern) and have evolved their contrasting 79 growth habits independently multiple times 7,20 . These Dune and Headland populations are 80 locally adapted [17][18][19][20] and their F2 hybrids have low fitness 21 , indicating the presence of 81 intrinsic reproductive isolation. Furthermore, performing genetic, physiological, and 82 ecological experimental studies is achievable in this system due to its short life cycle, diploid 83 inheritance, and small vegetative size. Therefore, the Senecio lautus species complex 84The physiological basis of repeated evolution in S. lautus 126Considering we identified a multitude of different auxin related genes between erect and 127 prostrate populations of S. lautus and the regulation and transport of auxin is well established 128 to modulate gravitropism in plants, we predicted that these divergent growth habits may be a 129 direct consequence of changes in the auxin pathway, and can therefore contribute to 130 d...

16Phenotypic plasticity can maintain population fitness in novel or changing environments if it allows the 17 phenotype to track the new trait optimum. Understanding how adaptation to contrasting environments 18 determines plastic responses can identify how plasticity evolves, and its potential to be adaptive in response 19 to environmental change. We sampled 79 genotypes from populations of two closely related but ecologically 20 divergent ragwort species (Senecio, Asteraceae), and transplanted multiple clones of each genotype into four 21 field sites along an elevational gradient representing each species' native range, the edge of their range, and 22 conditions outside their native range. At each transplant site, we quantified differences in survival, growth, 23 leaf morphology, chlorophyll fluorescence and gene expression for both species. Overall, the two species 24 differed in their sensitivity to the elevational gradient. As evidence of plasticity, leaf morphology changed 25 across the elevational gradient, with changes occurring in opposite directions for the two species. Differential 26 gene expression across the four field sites also revealed that the genetic pathways underlying plastic 27 responses were highly distinct in the two species. Despite the two species having diverged recently, 28 adaptation to contrasting habitats has resulted in the evolution of distinct sensitivities to environmental 29 variation, underlain by distinct forms of plasticity. 30 genotype-by-environment interactions, phenotypic plasticity, physiological plasticity, specialisation 32 33 62when the environment is predictable, leading to adaptive plasticity within the environmental limits 63 experienced during adaptation (Bradshaw 1965; Schlichting 1986; Baythavong and Stanton 2010). Whether 64 such plasticity will continue to be adaptive when exposed to novel conditions, such as those imposed by 65 4 climate change, remains an empirical issue (Ghalambor et al. 2007). Strong stabilising selection created by 66 predictable environments is expected to lead to specific plastic responses and reduce genetic variation for 67 plasticity (Oostra et al. 2018). By contrast, populations adapted to a wider range of habitats that are more 68 spatially and temporally variable are predicted to maintain genetic variation in plastic responses, increasing 69 the potential for selection on plasticity (Chevin et al. 2010). Detecting and characterising patterns of G×E for 70 a range of naturally occurring genotypes can help us understand whether evolutionary responses can occur 71 even if plasticity is constrained in certain directions (Via 1993; Chevin and Hoffmann 2017). 72 The genetic architecture underlying variation in plasticity is largely unknown (Fusco and Minelli 2010). 73 Plastic responses at the gene expression level are most likely controlled either by epiallelic control of the 74 genes themselves or allelic variation in the regulators of the genes (Rockman and Kruglyak 2006). If allelic 75 (sequence changes) or epiallelic (e.g. DNA me...