The hierarchical organization of autocatalytic reaction networks and its relevance to the origin of life.

Prior work on abiogenesis, the emergence of life from non-life, suggests that it requires chemical reaction networks that contain self-amplifying motifs, namely, autocatalytic cores. However, little is known about how the presence of multiple autocatalytic cores might allow for the gradual accretion of complexity on the path to life. To explore this problem, we develop the concept of a seed-dependent autocatalytic system (SDAS), which is a subnetwork that can autocatalytically self-maintain given a flux of food, but cannot be initiated by food alone. Rather, initiation of SDASs requires the transient introduction of chemical "seeds." We show that, depending on the topological relationship of SDASs in a chemical reaction network, a food-driven system can accrete complexity in a historically contingent manner, governed by rare seeding events. We develop new algorithms for detecting and analyzing SDASs in chemical reaction databases and describe parallels between multi-SDAS networks and biological ecosystems. Applying our algorithms to both an abiotic reaction network and a biochemical one, each driven by a set of simple food chemicals, we detect SDASs that are organized as trophic tiers, of which the higher tier can be seeded by relatively simple chemicals if the lower tier is already activated. This indicates that sequential activation of trophically organized SDASs by seed chemicals that are not much more complex than what already exist could be a mechanism of gradual complexification from relatively simple abiotic reactions to more complex life-like systems. Interestingly, in both reaction networks, higher-tier SDASs include chemicals that might alter emergent features of chemical systems and could serve as early targets of selection. Our analysis provides computational tools for analyzing very large chemical/biochemical reaction networks and suggests new approaches to studying abiogenesis in the lab.

Reticulate Evolution Helps Explain Apparent Homoplasy in Floral Biology and Pollination in Baobabs (Adansonia; Bombacoideae; Malvaceae).

Baobabs (Adansonia) are a cohesive group of tropical trees with a disjunct distribution in Australia, Madagascar, and continental Africa, and diverse flowers associated with two pollination modes. We used custom-targeted sequence capture in conjunction with new and existing phylogenetic comparative methods to explore the evolution of floral traits and pollination systems while allowing for reticulate evolution. Our analyses suggest that relationships in Adansonia are confounded by reticulation, with network inference methods supporting at least one reticulation event. The best supported hypothesis involves introgression between Adansonia rubrostipa and core Longitubae, both of which are hawkmoth pollinated with yellow/red flowers, but there is also some support for introgression between the African lineage and Malagasy Brevitubae, which are both mammal-pollinated with white flowers. New comparative methods for phylogenetic networks were developed that allow maximum-likelihood inference of ancestral states and were applied to study the apparent homoplasy in floral biology and pollination mode seen in Adansonia. This analysis supports a role for introgressive hybridization in morphological evolution even in a clade with highly divergent and geographically widespread species. Our new comparative methods for discrete traits on species networks are implemented in the software PhyloNetworks. [Comparative methods; Hyb-Seq; introgression; network inference; population trees; reticulate evolution; species tree inference; targeted sequence capture.].

An ecological framework for the analysis of prebiotic chemical reaction networks.

It is becoming widely accepted that very early in life's origin, even before the emergence of genetic encoding, reaction networks of diverse small chemicals might have manifested key properties of life, namely self-propagation and adaptive evolution. To explore this possibility, we formalize the dynamics of chemical reaction networks within the framework of chemical ecosystem ecology. To capture the idea that life-like chemical systems are maintained out of equilibrium by fluxes of energy-rich food chemicals, we model chemical ecosystems in well-mixed compartments that are subject to constant dilution by a solution with a fixed concentration of input chemicals. Modelling all chemical reactions as fully reversible, we show that seeding an autocatalytic cycle with tiny amounts of one or more of its member chemicals results in logistic growth of all member chemicals in the cycle. This finding justifies drawing an instructive analogy between an autocatalytic cycle and a biological species. We extend this finding to show that pairs of autocatalytic cycles can exhibit competitive, predator-prey, or mutualistic associations just like biological species. Furthermore, when there is stochasticity in the environment, particularly in the seeding of autocatalytic cycles, chemical ecosystems can show complex dynamics that can resemble evolution. The evolutionary character is especially clear when the network architecture results in ecological precedence, which makes a system's trajectory historically contingent on the order in which cycles are seeded. For all its simplicity, the framework developed here helps explain the onset of adaptive evolution in prebiotic chemical reaction networks, and can shed light on the origin of key biological attributes such as thermodynamic irreversibility and genetic encoding.

A Malvaceae mystery: A mallow maelstrom of genome multiplications and maybe misleading methods?

Previous research suggests that Gossypium has undergone a 5- to 6-fold multiplication following its divergence from Theobroma. However, the number of events, or where they occurred in the Malvaceae phylogeny remains unknown. We analyzed transcriptomic and genomic data from representatives of eight of the nine Malvaceae subfamilies. Phylogenetic analysis of nuclear data placed Dombeya (Dombeyoideae) as sister to the rest of Malvadendrina clade, but the plastid DNA tree strongly supported Durio (Helicteroideae) in this position. Intraspecific Ks plots indicated that all sampled taxa, except Theobroma (Byttnerioideae), Corchorus (Grewioideae), and Dombeya (Dombeyoideae), have experienced whole genome multiplications (WGMs). Quartet analysis suggested WGMs were shared by Malvoideae-Bombacoideae and Sterculioideae-Tilioideae, but did not resolve whether these are shared with each other or Helicteroideae (Durio). Gene tree reconciliation and Bayesian concordance analysis suggested a complex history. Alternative hypotheses are suggested, each involving two independent autotetraploid and one allopolyploid event. They differ in that one entails an allopolyploid origin for the Durio lineage, whereas the other invokes an allopolyploid origin for Malvoideae-Bombacoideae. We highlight the need for more genomic information in the Malvaceae and improved methods to resolve complex evolutionary histories that may include allopolyploidy, incomplete lineage sorting, and variable rates of gene and genome evolution.

Exclusivity offers a sound yet practical species criterion for bacteria despite abundant gene flow.

BACKGROUND: The question of whether bacterial species objectively exist has long divided microbiologists. A major source of contention stems from the fact that bacteria regularly engage in horizontal gene transfer (HGT), making it difficult to ascertain relatedness and draw boundaries between taxa. A natural way to define taxa is based on exclusivity of relatedness, which applies when members of a taxon are more closely related to each other than they are to any outsider. It is largely unknown whether exclusive bacterial taxa exist when averaging over the genome or are rare due to rampant hybridization. RESULTS: Here, we analyze a collection of 701 genomes representing a wide variety of environmental isolates from the family Streptomycetaceae, whose members are competent at HGT. We find that the presence/absence of auxiliary genes in the pan-genome displays a hierarchical (tree-like) structure that correlates significantly with the genealogy of the core-genome. Moreover, we identified the existence of many exclusive taxa, although individual genes often contradict these taxa. These conclusions were supported by repeating the analysis on 1,586 genomes belonging to the genus Bacillus. However, despite confirming the existence of exclusive groups (taxa), we were unable to identify an objective threshold at which to assign the rank of species. CONCLUSIONS: The existence of bacterial taxa is justified by considering average relatedness across the entire genome, as captured by exclusivity, but is rejected if one requires unanimous agreement of all parts of the genome. We propose using exclusivity to delimit taxa and conventional genome similarity thresholds to assign bacterial taxa to the species rank. This approach recognizes species that are phylogenetically meaningful, while also establishing some degree of comparability across species-ranked taxa in different bacterial clades.

The origin and early evolution of life in chemical composition space.

Life can be viewed as a localized chemical system that sits in the basin of attraction of a metastable dynamical attractor state that remains out of equilibrium with the environment. To explore the implications of this conception, I introduce an abstract coordinate system, chemical composition (CC Space), which summarizes the degree to which chemical systems are out of equilibrium with the bulk environment. A system's chemical disequilibrium (CD) is defined to be proportional to the Euclidean distance between the composition of a small region of physical space, a pixel, and the origin of CC space. Such a model implies that new living states arise through chance changes in local chemical concentration ("mutations") that cause chemical systems to move in CC space and enter the basin of attraction of a life state. The attractor of a life state comprises an autocatalytic set of chemicals whose essential ("keystone") species are produced at a higher rate than they are lost to the environment by diffusion, such that spatial growth of the life state is expected. This framework suggests that new life states are most likely to form at the interface between different physical phases, where the rate of diffusion of keystone species is tied to the low-diffusion regime, whereas food and waste products are subject to the more diffusive regime. Once life nucleates, for example on a mineral surface, it will tend to grow and generate variants as a result of additional mutations that find alternative life states. By jumping from life state to life state, systems can eventually occupy areas of CC space that are too far out of equilibrium with the environment to ever arise in a single mutational step. Furthermore, I propose that variation in the capacity of different surface associated life states to persist and compete may systematically favor states that have higher chemical disequilibrium. The model also suggests a simple and predictable path from surface-associated life to cell-like individuation. This dynamical systems theoretical framework provides an integrated view of the origin and early evolution of life and supports novel empirical approaches.

An Experimental Framework for Generating Evolvable Chemical Systems in the Laboratory.

Most experimental work on the origin of life has focused on either characterizing the chemical synthesis of particular biochemicals and their precursors or on designing simple chemical systems that manifest life-like properties such as self-propagation or adaptive evolution. Here we propose a new class of experiments, analogous to artificial ecosystem selection, where we select for spontaneously forming self-propagating chemical assemblages in the lab and then seek evidence of a response to that selection as a key indicator that life-like chemical systems have arisen. Since surfaces and surface metabolism likely played an important role in the origin of life, a key experimental challenge is to find conditions that foster nucleation and spread of chemical consortia on surfaces. We propose high-throughput screening of a diverse set of conditions in order to identify combinations of "food," energy sources, and mineral surfaces that foster the emergence of surface-associated chemical consortia that are capable of adaptive evolution. Identification of such systems would greatly advance our understanding of the emergence of self-propagating entities and the onset of adaptive evolution during the origin of life.

Long-term morphological stasis maintained by a plant-pollinator mutualism.

Many major branches in the Tree of Life are marked by stereotyped body plans that have been maintained over long periods of time. One possible explanation for this stasis is that there are genetic or developmental constraints that restrict the origin of novel body plans. An alternative is that basic body plans are potentially quite labile, but are actively maintained by natural selection. We present evidence that the conserved floral morphology of a species-rich flowering plant clade, Malpighiaceae, has been actively maintained for tens of millions of years via stabilizing selection imposed by their specialist New World oil-bee pollinators. Nine clades that have lost their primary oil-bee pollinators show major evolutionary shifts in specific floral traits associated with oil-bee pollination, demonstrating that developmental constraint is not the primary cause of morphological stasis in Malpighiaceae. Interestingly, Malpighiaceae show a burst in species diversification coinciding with the origin of this plant-pollinator mutualism. One hypothesis to account for radiation despite morphological stasis is that although selection on pollinator efficiency explains the origin of this unique and conserved floral morphology, tight pollinator specificity subsequently permitted greatly enhanced diversification in this system.

Whole genome duplication in coast redwood (Sequoia sempervirens) and its implications for explaining the rarity of polyploidy in conifers.

Polyploidy is common and an important evolutionary factor in most land plant lineages, but it is rare in gymnosperms. Coast redwood (Sequoia sempervirens) is one of just two polyploid conifer species and the only hexaploid. Evidence from fossil guard cell size suggests that polyploidy in Sequoia dates to the Eocene. Numerous hypotheses about the mechanism of polyploidy and parental genome donors have been proposed, based primarily on morphological and cytological data, but it remains unclear how Sequoia became polyploid and why this lineage overcame an apparent gymnosperm barrier to whole-genome duplication (WGD). We sequenced transcriptomes and used phylogenetic inference, Bayesian concordance analysis and paralog age distributions to resolve relationships among gene copies in hexaploid coast redwood and close relatives. Our data show that hexaploidy in coast redwood is best explained by autopolyploidy or, if there was allopolyploidy, it happened within the Californian redwood clade. We found that duplicate genes have more similar sequences than expected, given the age of the inferred polyploidization. Conflict between molecular and fossil estimates of WGD can be explained if diploidization occurred very slowly following polyploidization. We extrapolate from this to suggest that the rarity of polyploidy in gymnosperms may be due to slow diploidization in this clade.

Clade-specific positive selection on a developmental gene: BRANCHLESS TRICHOME and the evolution of stellate trichomes in Physaria (Brassicaceae).

Positive selection is known to drive the evolution of genes involved in evolutionary arms races, but what role does it play in the evolution of genes involved in developmental processes? We used the single-celled epidermal trichomes of Brassicaceae as a model to uncover the molecular evolutionary processes that contributed to the transition from dendritic trichomes, as seen in most species of Brassicaceae, to the distinctive stellate trichomes of the genus Physaria. We explored the role of positive selection on the evolution of BRANCHLESS TRICHOME (BLT), a candidate gene for changes in trichome branching pattern. Maximum likelihood models of codon evolution point to a shift in selective pressure affecting the evolution of BLT across the entire Physaria clade, and we found strong evidence that positive selection has acted on a subset of Physaria BLT codons. Almost all of the 10 codon sites with the highest probability of having evolved under positive selection are clustered in a predicted coiled-coil domain, pointing to changes in protein-protein interactions. Thus, our findings suggest that selection acted on BLT to modify its interactions with other proteins. The fact that positive selection occurred throughout the radiation of Physaria could reflect selection to stabilize development in response to an abrupt switch from the dendritic form to the stellate form, divergent selection for diversification of the stellate form, or both. These results point to the need for evolutionary developmental studies of BLT and its interacting proteins in Physaria.

Revisiting the phylogeny of Bombacoideae (Malvaceae): Novel relationships, morphologically cohesive clades, and a new tribal classification based on multilocus phylogenetic analyses.

Bombacoideae (Malvaceae) is a clade of deciduous trees with a marked dominance in many forests, especially in the Neotropics. The historical lack of a well-resolved phylogenetic framework for Bombacoideae hinders studies in this ecologically important group. We reexamined phylogenetic relationships in this clade based on a matrix of 6465 nuclear (ETS, ITS) and plastid (matK, trnL-trnF, trnS-trnG) DNA characters. We used maximum parsimony, maximum likelihood, and Bayesian inference to infer relationships among 108 species (∼70% of the total number of known species). We analyzed the evolution of selected morphological traits: trunk or branch prickles, calyx shape, endocarp type, seed shape, and seed number per fruit, using ML reconstructions of their ancestral states to identify possible synapomorphies for major clades. Novel phylogenetic relationships emerged from our analyses, including three major lineages marked by fruit or seed traits: the winged-seed clade (Bernoullia, Gyranthera, and Huberodendron), the spongy endocarp clade (Adansonia, Aguiaria, Catostemma, Cavanillesia, and Scleronema), and the Kapok clade (Bombax, Ceiba, Eriotheca, Neobuchia, Pachira, Pseudobombax, Rhodognaphalon, and Spirotheca). The Kapok clade, the most diverse lineage of the subfamily, includes sister relationships (i) between Pseudobombax and "Pochota fendleri" a historically incertae sedis taxon, and (ii) between the Paleotropical genera Bombax and Rhodognaphalon, implying just two bombacoid dispersals to the Old World, the other one involving Adansonia. This new phylogenetic framework offers new insights and a promising avenue for further evolutionary studies. In view of this information, we present a new tribal classification of the subfamily, accompanied by an identification key.

Exploring Tree-Like and Non-Tree-Like Patterns Using Genome Sequences: An Example Using the Inbreeding Plant Species Arabidopsis thaliana (L.) Heynh.

Genome sequence data contain abundant information about genealogical history, but methods for extracting and interpreting this information are not yet fully developed. We analyzed genome sequences for multiple accessions of the selfing plant, Arabidopsis thaliana, with the goal of better understanding its genealogical history. As expected from accessions of the same species, we found much discordance between nuclear gene trees. Nonetheless, we inferred the optimal population tree under the assumption that all discordance is due to incomplete lineage sorting. To cope with the size of the data (many genes and many taxa), our pipeline is based on parallel computing and divides the problem into four-taxon trees. However, just because a population tree can be estimated does not mean that the assumptions of the multispecies coalescent model hold. Therefore, we implemented a new, nonparametric test to evaluate whether a population tree adequately explains the observed quartet frequencies (the frequencies of gene trees with each resolution of each four-taxon set). This test also considers other models: panmixia and a partially resolved population tree, that is, a tree in which some nodes are collapsed into local panmixia. We found that a partially resolved population tree provides the best fit to the data, providing evidence for tree-like structure within A. thaliana, qualitatively similar to what might be expected between different, closely related species. Further, we show that the pattern of deviation from expectations can be used to identify instances of introgression and detect one clear case of reticulation among ecotypes that have come into contact in the United Kingdom. Our study illustrates how we can use genome sequence data to evaluate whether phylogenetic relationships are strictly tree-like or reticulating.

A comparison of autogenous theories for the origin of eukaryotic cells.

PREMISE: Eukaryotic cells have many unique features that all evolved on the stem lineage of living eukaryotes, making it difficult to reconstruct the order in which they accumulated. Nuclear endosymbiotic theories hold that three prokaryotes (nucleus, cytoplasm, and mitochondrion) came together to form a eukaryotic cell, whereas autogenous models hold that the nucleus and cytoplasm formed through evolutionary changes in a single prokaryotic lineage. Given several problems with nuclear endosymbiotic theories, this review focuses on autogenous models. KEY INSIGHTS: Until recently all autogenous models assumed an outside-in (OI) topology, proposing that the nuclear envelope was formed from membrane-bound vesicles within the original cell body. Buzz Baum and I recently proposed an inside-out (IO) alternative, suggesting that the nucleus corresponds to the original cell body, with the cytoplasmic compartment deriving from extracellular protrusions. In this review, I show that OI and IO models are compatible with both mitochondria early (ME) or mitochondria late (ML) formulations. Whereas ME models allow that the relationship between mitochondria and host was mutualistic from the outset, ML models imply that the association began with predation or parasitism, becoming mutualistic later. In either case, the mutualistic interaction that eventually formed was probably syntrophic. CONCLUSIONS: Diverse features of eukaryotic cell biology align well with the IOME model, but it would be premature to rule out the OIME model. ML models require that phagocytosis, a complex and energy expensive process, evolved before mitochondria, which seems unlikely. Nonetheless, further research is needed, especially resolution of the phylogenetic affinities of mitochondria.

An inside-out origin for the eukaryotic cell.

BACKGROUND: Although the origin of the eukaryotic cell has long been recognized as the single most profound change in cellular organization during the evolution of life on earth, this transition remains poorly understood. Models have always assumed that the nucleus and endomembrane system evolved within the cytoplasm of a prokaryotic cell. RESULTS: Drawing on diverse aspects of cell biology and phylogenetic data, we invert the traditional interpretation of eukaryotic cell evolution. We propose that an ancestral prokaryotic cell, homologous to the modern-day nucleus, extruded membrane-bound blebs beyond its cell wall. These blebs functioned to facilitate material exchange with ectosymbiotic proto-mitochondria. The cytoplasm was then formed through the expansion of blebs around proto-mitochondria, with continuous spaces between the blebs giving rise to the endoplasmic reticulum, which later evolved into the eukaryotic secretory system. Further bleb-fusion steps yielded a continuous plasma membrane, which served to isolate the endoplasmic reticulum from the environment. CONCLUSIONS: The inside-out theory is consistent with diverse kinds of data and provides an alternative framework by which to explore and understand the dynamic organization of modern eukaryotic cells. It also helps to explain a number of previously enigmatic features of cell biology, including the autonomy of nuclei in syncytia and the subcellular localization of protein N-glycosylation, and makes many predictions, including a novel mechanism of interphase nuclear pore insertion.

Evidence of Heritability in Prebiotically Realistic Membrane-Bound Systems.

The vesicles of short chain amphiphiles have been demonstrated to grow and divide. Here, we explored whether vesicle populations show evidence of heritability. We prepared 1:1 decanoic acid:decylamine vesicles with or without a detergent and in either water or prebiotic soup, a mixture of compounds that might have been present on early Earth. The mixtures were subjected to transfer with dilution, where, after 24 h of incubation (one generation), we transferred 10% of the mix into a 90% volume of a fresh vesicle-containing solution. This was continued for 30 generations. Samples with a history of transfers were compared to no-transfer controls (NTCs), initiated each generation using the same solutions but without 10% of the prior generation. We compared the vesicle size distribution and chemical composition of the transfer samples and NTCs and compared their fluorescence signals in the presence of Nile Red dye. We observe changes in the vesicle size but did not detect differences in the chemical composition. In the samples with detergent and soup, we observed irregular changes in the Nile Red fluorescence, with a tendency for parent and offspring samples to have correlated values, suggestive of heritability. This last result, combined with evidence of temporal autocorrelation across generations, suggests the possibility that vesicles could respond to selection.

Precise spatio-temporal regulation of the anthocyanin biosynthetic pathway leads to petal spot formation in Clarkia gracilis (Onagraceae).

Petal spots are widespread in angiosperms and are often implicated in pollinator attraction. Clarkia gracilis petals each have a single red-purple spot that contrasts against a pink background. The position and presence of spots in C. gracilis are determined by the epistatic interaction of alleles at two as yet unidentified loci. We used HPLC to identify the different pigments produced in the petals, and qualitative and quantitative RT-PCR to assay for spatio-temporal patterns of expression of different anthocyanin pathway genes. We found that spots contain different pigments from the remainder of the petal, being composed of cyanidin/peonidin-based, instead of malvidin-based anthocyanins. Expression assays of anthocyanin pathway genes showed that the dihydroflavonol-4-reductase 2 (Dfr2) gene has a spot-specific expression pattern and acts as a switch for spot production. Co-segregation analyses implicated the gene products of the P and I loci as trans-regulators of this switch. Spot pigments appear earlier in development as a result of early expression of Dfr2 and the flavonoid 3' hydroxylase 1 (F3'h1) gene. Pigments in the background appear later, as a result of later expression of Dfr1 and the flavonoid 3'-5' hydroxylase 1 (F3'5'h1) genes. The evolution of this spot production mechanism appears to have been facilitated by duplication of the Dfr gene and to have required substantial reworking of the anthocyanin pathway regulatory network.

The ecology-evolution continuum and the origin of life.

Prior research on evolutionary mechanisms during the origin of life has mainly assumed the existence of populations of discrete entities with information encoded in genetic polymers. Recent theoretical advances in autocatalytic chemical ecology establish a broader evolutionary framework that allows for adaptive complexification prior to the emergence of bounded individuals or genetic encoding. This framework establishes the formal equivalence of cells, ecosystems and certain localized chemical reaction systems as autocatalytic chemical ecosystems (ACEs): food-driven (open) systems that can grow due to the action of autocatalytic cycles (ACs). When ACEs are organized in meta-ecosystems, whether they be populations of cells or sets of chemically similar environmental patches, evolution, defined as change in AC frequency over time, can occur. In cases where ACs are enriched because they enhance ACE persistence or dispersal ability, evolution is adaptive and can build complexity. In particular, adaptive evolution can explain the emergence of self-bounded units (e.g. protocells) and genetic inheritance mechanisms. Recognizing the continuity between ecological and evolutionary change through the lens of autocatalytic chemical ecology suggests that the origin of life should be seen as a general and predictable outcome of driven chemical ecosystems rather than a phenomenon requiring specific, rare conditions.

The need for molecular genetic perspectives in evolutionary education (and vice versa).

Despite evolution being the central concept of biology, many biology students do not understand the core principles of evolutionary theory. Here, we propose that integrating evolution throughout the biology curriculum, and incorporating molecular biology and molecular genetic perspectives, will help students not only to achieve a better understanding of evolution, but also to appreciate how modern evolutionary biology research is practiced. However, realizing this vision will require changes in the way that biology is taught. Educators will need to teach from a broadened perspective and to utilize new strategies for maximizing student learning. However, if this vision is realized then strides can be made in advancing the public understanding of evolution.

The Caribbean slipper spurge Euphorbia tithymaloides: the first example of a ring species in plants.

A ring species arises when a parental population expands around an area of unsuitable habitat in such a way that when the two fronts meet they behave as distinct species while still being connected through a series of intergrading populations. Ring species offer great possibilities for studying the forces causing species divergence (e.g. the nature of pre-zygotic or post-zygotic reproductive isolation) or helping to maintain species integrity (e.g. reinforcement). Yet, ring species are extremely rare, and have only been documented convincingly in animals. Here, we present phylogenetic analyses of two nuclear gene regions from the Caribbean slipper spurge (Euphorbia tithymaloides) species complex that provide evidence that this group forms a ring species. These data show that the species complex originated in the area where Mexico and Guatemala meet, and expanded around the Caribbean basin along two distinct fronts: one eastward through the Yucatan Peninsula and into the Greater Antilles (GA); one southeastward through northern South America and then northward to the Lesser Antilles and eastern GA. The two terminal forms co-occur in the Virgin Islands and appear to be morphologically and ecologically distinct. Thus, our results suggest that Euphorbia tithymaloides is the first compelling example of a ring species in plants.