Introgression is widespread in the radiation of carnivorous Nepenthes pitcher plants.

Introgression and hybridization are important processes in plant evolution, but they are difficult to study from a phylogenetic perspective, because they conflict with the bifurcating evolutionary history typically depicted in phylogenetic models. The role of hybridization in plant evolution is best documented in the form of allo-polyploidizations. In contrast, homoploid hybridization and introgression are less explored, although they may be crucial in adaptive radiations. Here we employ genome-wide data (ddRAD-seq, transcriptomes) to investigate the evolutionary history of Nepenthes, a radiation of c. 160 species of iconic carnivorous plants mainly from tropical Asia. Our data indicates that the main radiation is only c. 5 million years old, and confirms previous bifurcating phylogenies. However, due to a greatly expanded number of loci, we were able test for the first time the long-standing hypotheses of introgression and historical hybridization. The genus presents one very clear case of organellar capture between two distantly related but sympatric groups. Furthermore, all Nepenthes species show introgression signals in their nuclear genomes, as uncovered by a general survey of ABBA-BABA-like statistics. The ancestor of the rapid main radiation shows ancestry from two deeply diverged lineages, as indicated by phylogenetic network analyses. All major clades of the main radiation show further introgression both within and between each other, as suggested by admixture graphs. Our study supports the hypothesis that rapid adaptive radiations are hotspots of introgression in the tree of life, and highlights the need to consider non-treelike processes in evolutionary studies of Nepenthes in particular.

'Insect aquaplaning' on a superhydrophilic hairy surface: how Heliamphora nutans Benth. pitcher plants capture prey.

Trichomes are a common feature of plants and perform important and diverse functions. Here, we show that the inward-pointing hairs on the inner wall of insect-trapping Heliamphora nutans pitchers are highly wettable, causing water droplets to spread rapidly across the surface. Wetting strongly enhanced the slipperiness and increased the capture rate for ants from 29 to 88 per cent. Force measurements and tarsal ablation experiments revealed that wetting affected the insects' adhesive pads but not the claws, similar to the 'aquaplaning' mechanism of (unrelated) Asian Nepenthes pitcher plants. The inward-pointing trichomes provided much higher traction when insects were pulled outwards. The wetness-dependent capture mechanisms of H. nutans and Nepenthes pitchers present a striking case of functional convergence, whereas the use of wettable trichomes constitutes a previously unknown mechanism to make plant surfaces slippery.

Subgenome dominance shapes novel gene evolution in the decaploid pitcher plant Nepenthes gracilis.

Subgenome dominance after whole-genome duplication generates distinction in gene number and expression at the level of chromosome sets, but it remains unclear how this process may be involved in evolutionary novelty. Here we generated a chromosome-scale genome assembly of the Asian pitcher plant Nepenthes gracilis to analyse how its novel traits (dioecy and carnivorous pitcher leaves) are linked to genomic evolution. We found a decaploid karyotype and a clear indication of subgenome dominance. A male-linked and pericentromerically located region on the putative sex chromosome was identified in a recessive subgenome and was found to harbour three transcription factors involved in flower and pollen development, including a likely neofunctionalized LEAFY duplicate. Transcriptomic and syntenic analyses of carnivory-related genes suggested that the paleopolyploidization events seeded genes that subsequently formed tandem clusters in recessive subgenomes with specific expression in the digestive zone of the pitcher, where specialized cells digest prey and absorb derived nutrients. A genome-scale analysis suggested that subgenome dominance likely contributed to evolutionary innovation by permitting recessive subgenomes to diversify functions of novel tissue-specific duplicates. Our results provide insight into how polyploidy can give rise to novel traits in divergent and successful high-ploidy lineages.

High rates of evolution preceded shifts to sex-biased gene expression in Leucadendron, the most sexually dimorphic angiosperms.

Differences between males and females are usually more subtle in dioecious plants than animals, but strong sexual dimorphism has evolved convergently in the South African Cape plant genus Leucadendron. Such sexual dimorphism in leaf size is expected largely to be due to differential gene expression between the sexes. We compared patterns of gene expression in leaves among 10 Leucadendron species across the genus. Surprisingly, we found no positive association between sexual dimorphism in morphology and the number or the percentage of sex-biased genes (SBGs). Sex bias in most SBGs evolved recently and was species specific. We compared rates of evolutionary change in expression for genes that were sex biased in one species but unbiased in others and found that SBGs evolved faster in expression than unbiased genes. This greater rate of expression evolution of SBGs, also documented in animals, might suggest the possible role of sexual selection in the evolution of gene expression. However, our comparative analysis clearly indicates that the more rapid rate of expression evolution of SBGs predated the origin of bias, and shifts towards bias were depleted in signatures of adaptation. Our results are thus more consistent with the view that sex bias is simply freer to evolve in genes less subject to constraints in expression level.

A neutral model for the loss of recombination on sex chromosomes.

The loss of recombination between sex chromosomes has occurred repeatedly throughout nature, with important implications for their subsequent evolution. Explanations for this remarkable convergence have generally invoked only adaptive processes (e.g. sexually antagonistic selection); however, there is still little evidence for these hypotheses. Here we propose a model in which recombination on sex chromosomes is lost due to the neutral accumulation of sequence divergence adjacent to (and thus, in linkage disequilibrium with) the sex determiner. Importantly, we include in our model the fact that sequence divergence, in any form, reduces the probability of recombination between any two sequences. Using simulations, we show that, under certain conditions, a region of suppressed recombination arises and expands outwards from the sex-determining locus, under purely neutral processes. Further, we show that the rate and pattern of recombination loss are sensitive to the pre-existing recombination landscape of the genome and to sex differences in recombination rates, with patterns consistent with evolutionary strata emerging under some conditions. We discuss the applicability of these results to natural systems. This article is part of the theme issue 'Challenging the paradigm in sex chromosome evolution: empirical and theoretical insights with a focus on vertebrates (Part I)'.

Population Genomics of the "Arcanum" Species Group in Wild Tomatoes: Evidence for Separate Origins of Two Self-Compatible Lineages.

Given their diverse mating systems and recent divergence, wild tomatoes (Solanum section Lycopersicon) have become an attractive model system to study ecological divergence, the build-up of reproductive barriers, and the causes and consequences of the breakdown of self-incompatibility. Here we report on a lesser-studied group of species known as the "Arcanum" group, comprising the nominal species Solanum arcanum, Solanum chmielewskii, and Solanum neorickii. The latter two taxa are self-compatible but are thought to self-fertilize at different rates, given their distinct manifestations of the morphological "selfing syndrome." Based on experimental crossings and transcriptome sequencing of a total of 39 different genotypes from as many accessions representing each species' geographic range, we provide compelling evidence for deep genealogical divisions within S. arcanum; only the self-incompatible lineage known as "var. marañón" has close genealogical ties to the two self-compatible species. Moreover, there is evidence under multiple inference schemes for different geographic subsets of S. arcanum var. marañón being closest to S. chmielewskii and S. neorickii, respectively. To broadly characterize the population-genomic consequences of these recent mating-system transitions and their associated speciation events, we fit demographic models indicating strong reductions in effective population size, congruent with reduced nucleotide and S-locus diversity in the two independently derived self-compatible species.

Evolutionary footprints of a cold relic in a rapidly warming world.

With accelerating global warming, understanding the evolutionary dynamics of plant adaptation to environmental change is increasingly urgent. Here, we reveal the enigmatic history of the genus Cochlearia (Brassicaceae), a Pleistocene relic that originated from a drought-adapted Mediterranean sister genus during the Miocene. Cochlearia rapidly diversified and adapted to circum-Arctic regions and other cold-characterized habitat types during the Pleistocene. This sudden change in ecological preferences was accompanied by a highly complex, reticulate polyploid evolution, which was apparently triggered by the impact of repeated Pleistocene glaciation cycles. Our results illustrate that two early diversified Arctic-alpine diploid gene pools contributed differently to the evolution of this young polyploid genus now captured in a cold-adapted niche. Metabolomics revealed central carbon metabolism responses to cold in diverse species and ecotypes, likely due to continuous connections to cold habitats that may have facilitated widespread adaptation to alpine and subalpine habitats, and which we speculate were coopted from existing drought adaptations. Given the growing scientific interest in the adaptive evolution of temperature-related traits, our results provide much-needed taxonomic and phylogenomic resolution of a model system as well as first insights into the origins of its adaptation to cold.

Identification of sex-linked markers in the sexually cryptic coco de mer: are males and females produced in equal proportions?

Lodoicea maldivica (coco de mer) is a long-lived dioecious palm in which male and female plants are visually indistinguishable when immature, only becoming sexually dimorphic as adults, which in natural forest can take as much as 50 years. Most adult populations in the Seychelles exhibit biased sex ratios, but it is unknown whether this is due to different proportions of male and female plants being produced or to differential mortality. In this study, we developed sex-linked markers in Lodoicea using ddRAD sequencing, enabling us to reliably determine the gender of immature individuals. We screened 589 immature individuals to explore sex ratios across life stages in Lodoicea. The two sex-specific markers resulted in the amplification of male-specific bands (Lm123977 at 405 bp and Lm435135 at 130 bp). Our study of four sub-populations of Lodoicea on the islands of Praslin and Curieuse revealed that the two sexes were produced in approximately equal numbers, with no significant deviation from a 1:1 ratio before the adult stage. We conclude that sex in Lodoicea is genetically determined, suggesting that Lodoicea has a chromosomal sex determination system in which males are the heterogametic sex (XY) and females are homogametic (XX). We discuss the potential causes for observed biased sex ratios in adult populations, and the implications of our results for the life history, ecology and conservation management of Lodoicea.

Sex is determined by XY chromosomes across the radiation of dioecious Nepenthes pitcher plants.

Species with separate sexes (dioecy) are a minority among flowering plants, but dioecy has evolved multiple times independently in their history. The sex-determination system and sex-linked genomic regions are currently identified in a limited number of dioecious plants only. Here, we study the sex-determination system in a genus of dioecious plants that lack heteromorphic sex chromosomes and are not amenable to controlled breeding: Nepenthes pitcher plants. We genotyped wild populations of flowering males and females of three Nepenthes taxa using ddRAD-seq and sequenced a male inflorescence transcriptome. We developed a statistical tool (privacy rarefaction) to distinguish true sex specificity from stochastic noise in read coverage of sequencing data from wild populations and identified male-specific loci and XY-patterned single nucleotide polymorphsims (SNPs) in all three Nepenthes taxa, suggesting the presence of homomorphic XY sex chromosomes. The male-specific region of the Y chromosome showed little conservation among the three taxa, except for the essential pollen development gene DYT1 that was confirmed as male specific by PCR in additional Nepenthes taxa. Hence, dioecy and part of the male-specific region of the Nepenthes Y-chromosomes likely have a single evolutionary origin.

A novel type of nutritional ant-plant interaction: ant partners of carnivorous pitcher plants prevent nutrient export by dipteran pitcher infauna.

Many plants combat herbivore and pathogen attack indirectly by attracting predators of their herbivores. Here we describe a novel type of insect-plant interaction where a carnivorous plant uses such an indirect defence to prevent nutrient loss to kleptoparasites. The ant Camponotus schmitzi is an obligate inhabitant of the carnivorous pitcher plant Nepenthes bicalcarata in Borneo. It has recently been suggested that this ant-plant interaction is a nutritional mutualism, but the detailed mechanisms and the origin of the ant-derived nutrient supply have remained unexplained. We confirm that N. bicalcarata host plant leaves naturally have an elevated (15)N/(14)N stable isotope abundance ratio (δ(15)N) when colonised by C. schmitzi. This indicates that a higher proportion of the plants' nitrogen is insect-derived when C. schmitzi ants are present (ca. 100%, vs. 77% in uncolonised plants) and that more nitrogen is available to them. We demonstrated direct flux of nutrients from the ants to the host plant in a (15)N pulse-chase experiment. As C. schmitzi ants only feed on nectar and pitcher contents of their host, the elevated foliar δ(15)N cannot be explained by classic ant-feeding (myrmecotrophy) but must originate from a higher efficiency of the pitcher traps. We discovered that C. schmitzi ants not only increase the pitchers' capture efficiency by keeping the pitchers' trapping surfaces clean, but they also reduce nutrient loss from the pitchers by predating dipteran pitcher inhabitants (infauna). Consequently, nutrients the pitchers would have otherwise lost via emerging flies become available as ant colony waste. The plants' prey is therefore conserved by the ants. The interaction between C. schmitzi, N. bicalcarata and dipteran pitcher infauna represents a new type of mutualism where animals mitigate the damage by nutrient thieves to a plant.