Evolutionary signatures of a trade-off in direct and indirect defenses across the wild grape genus, Vitis.

Evolutionary correlations between chemical defense and protection by mutualist bodyguards have been long predicted, but tests of these patterns remain rare. We use a phylogenetic framework to test for evolutionary correlations indicative of trade-offs or synergisms between direct defense in the form of plant secondary metabolism and indirect defense in the form of leaf domatia, across 33 species in the wild grape genus, Vitis. We also performed a bioassay with a generalist herbivore to associate our chemical phenotypes with herbivore palatability. Finally, we tested whether defensive traits correlated with the average abiotic characteristics of each species' contemporary range and whether these correlations were consistent with plant defense theory. We found a negative evolutionary correlation between domatia size and the diversity of secondary metabolites in Vitis leaf tissue across the genus, and also that leaves with a higher diversity and richness of secondary metabolites were less palatable to a generalist herbivore, consistent with a trade-off in chemical and mutualistic defense investment. Predictions from plant defense theory were not supported by associations between investment in defense phenotypes and abiotic variables. Our work demonstrates an evolutionary pattern indicative of a trade-off between indirect and direct defense strategies across the Vitis genus.

Macroevolution of protective coloration across caterpillars reflects relationships with host plants.

A critical function of animal coloration is avoiding attack, either by warning predators or reducing detectability. Evolution of these divergent strategies may depend on prey palatability and apparency to predators: conspicuous coloration may be favoured if species are distasteful, or habitats make hiding difficult; by contrast, camouflage may be effective if prey lack defences or environments are visually complex. For insect herbivores, host plants provide both chemical defence and the background against which they are detected or obscured; thus, plant traits may be key to coloration in these foundational terrestrial organisms. We use 1808 species of larval Lepidoptera to explore macroevolution of protective coloration strategy. We find that colour and pattern evolve jointly in caterpillars, similar to an array of species across the animal kingdom, while individual elements of coloration evolve closely with diet ecology. Consistent with key tenets of plant defence and plant-herbivore coevolutionary theory, conspicuous colours are associated with herbaceous host plants-thought to be defended by toxins-while camouflage colours and patterns are associated with woody plants and grasses. Contrary to theory, dietary specialization is not associated with conspicuous coloration. Our results add valuable insights into the evolutionary forces shaping colour and pattern in nature.

Modeling the Evolution of Rates of Continuous Trait Evolution.

Rates of phenotypic evolution vary markedly across the tree of life, from the accelerated evolution apparent in adaptive radiations to the remarkable evolutionary stasis exhibited by so-called "living fossils." Such rate variation has important consequences for large-scale evolutionary dynamics, generating vast disparities in phenotypic diversity across space, time, and taxa. Despite this, most methods for estimating trait evolution rates assume rates vary deterministically with respect to some variable of interest or change infrequently during a clade's history. These assumptions may cause underfitting of trait evolution models and mislead hypothesis testing. Here, we develop a new trait evolution model that allows rates to vary gradually and stochastically across a clade. Further, we extend this model to accommodate generally decreasing or increasing rates over time, allowing for flexible modeling of "early/late bursts" of trait evolution. We implement a Bayesian method, termed "evolving rates" (evorates for short), to efficiently fit this model to comparative data. Through simulation, we demonstrate that evorates can reliably infer both how and in which lineages trait evolution rates varied during a clade's history. We apply this method to body size evolution in cetaceans, recovering substantial support for an overall slowdown in body size evolution over time with recent bursts among some oceanic dolphins and relative stasis among beaked whales of the genus Mesoplodon. These results unify and expand on previous research, demonstrating the empirical utility of evorates. [cetacea; macroevolution; comparative methods; phenotypic diversity; disparity; early burst; late burst].

Six Principles for Embracing Gender and Sexual Diversity in Postsecondary Biology Classrooms.

Sexual and gender minorities face considerable inequities in society, including in science. In biology, course content provides opportunities to challenge harmful preconceptions about what is "natural" while avoiding the notion that anything found in nature is inherently good (the appeal-to-nature fallacy). We provide six principles for instructors to teach sex- and gender-related topics in postsecondary biology in a more inclusive and accurate manner: highlighting biological diversity early, presenting the social and historical context of science, using inclusive language, teaching the iterative process of science, presenting students with a diversity of role models, and developing a classroom culture of respect and inclusion. To illustrate these six principles, we review the many definitions of sex and demonstrate applying the principles to three example topics: sexual reproduction, sex determination or differentiation, and sexual selection. These principles provide a tangible starting place to create more scientifically accurate, engaging, and inclusive classrooms.

Generalising indirect defence and resistance of plants.

Indirect defence, the adaptive top-down control of herbivores by plant traits that enhance predation, is a central component of plant-herbivore interactions. However, the scope of interactions that comprise indirect defence and associated ecological and evolutionary processes has not been clearly defined. We argue that the range of plant traits that mediate indirect defence is much greater than previously thought, and we further organise major concepts surrounding their ecological functioning. Despite the wide range of plant traits and interacting organisms involved, indirect defences show commonalities when grouped. These categories are based on whether indirect defences boost natural enemy abundance via food or shelter resources, or, alternatively, increase natural enemy foraging efficiency via information or alteration of habitat complexity. The benefits of indirect defences to natural enemies should be further explored to establish the conditions in which indirect defence generates a plant-natural enemy mutualism. By considering the broader scope of plant-herbivore-natural enemy interactions that comprise indirect defence, we can better understand plant-based food webs, as well as the evolutionary processes that have shaped them.

Toxicity of the spiny thick-foot Pachypodium.

PREMISE OF THE STUDY: Pachypodium (Apocynaceae) is a genus of iconic stem-succulent and poisonous plants endemic to Madagascar and southern Africa. We tested hypotheses about the mode of action and macroevolution of toxicity in this group. We further hypothesized that while monarch butterflies are highly resistant to cardenolide toxins (a type of cardiac glycoside) from American Asclepias, they may be negatively affected by Pachypodium defenses, which evolved independently. METHODS: We grew 16 of 21 known Pachypodium spp. and quantified putative cardenolides by HPLC and also by inhibition of animal Na+ /K+ -ATPase (the physiological target of cardiac glycosides) using an in vitro assay. Pachypodium extracts were tested against monarch caterpillars in a feeding bioassay. We also tested four Asclepias spp. and five Pachypodium spp. extracts, contrasting inhibition of the cardenolide-sensitive porcine Na+ /K+ -ATPase to the monarch's resistant form. KEY RESULTS: We found evidence for low cardenolides by HPLC, but substantial toxicity when extracts were assayed on Na+ /K+ -ATPases. Toxicity showed phylogenetic signal, and taller species showed greater toxicity (this was marginal after phylogenetic correction). Application of Pachypodium extracts to milkweed leaves reduced monarch growth, and this was predicted by inhibition of the sensitive Na+ /K+ -ATPase in phylogenetic analyses. Asclepias extracts were 100-fold less potent against the monarch compared to the porcine Na+ /K+ -ATPase, but this difference was absent for Pachypodium extracts. CONCLUSIONS: Pachypodium contains potent toxicity capable of inhibiting sensitive and cardenolide-adapted Na+ /K+ -ATPases. Given the monarch's sensitivity to Pachypodium, we suggest that these plants contain novel cardiac glycosides or other compounds that facilitate toxicity by binding to Na+ /K+ -ATPases.

The evolution of floral signals in relation to range overlap in a clade of California Jewelflowers (Streptanthus s.l.).

Because of their function as reproductive signals in plants, floral traits experience distinct selective pressures related to their role in speciation, reinforcement, and prolonged coexistence with close relatives. However, few studies have investigated whether population-level processes translate into detectable signatures at the macroevolutionary scale. Here, we ask whether patterns of floral trait evolution and range overlap across a clade of California Jewelflowers reflect processes hypothesized to shape floral signal differentiation at the population level. We found a pattern of divergence in floral scent composition across the clade such that close relatives had highly disparate floral scents given their age. Accounting for range overlap with close relatives explained additional variation in floral scent over time, with sympatric species pairs having diverged more than allopatric species pairs given their age. However, three other floral traits (flower size, scent complexity and flower color) did not fit these patterns, failing to deviate from a null Brownian motion model of evolution. Together, our results suggest that selection for divergence among close relatives in the composition of floral scents may play a key, sustained role in mediating speciation and coexistence dynamics across this group, and that signatures of these dynamics may persist at the macroevolutionary scale.

Symbioses with nitrogen-fixing bacteria: nodulation and phylogenetic data across legume genera.

How species interactions shape global biodiversity and influence diversification is a central - but also data-hungry - question in evolutionary ecology. Microbially based mutualisms are widespread and could cause diversification by ameliorating stress and thus allowing organisms to colonize and adapt to otherwise unsuitable habitats. Yet the role of these interactions in generating species diversity has received limited attention, especially across large taxonomic groups. In the massive angiosperm family Leguminosae, plants often associate with root-nodulating bacteria that ameliorate nutrient stress by fixing atmospheric nitrogen. These symbioses are ecologically-important interactions, influencing community assembly, diversity, and succession, contributing ~100-290 million tons of N annually to natural ecosystems, and enhancing growth of agronomically-important forage and crop plants worldwide. In recent work attempting to determine whether mutualism with N-fixing bacteria led to increased diversification across legumes, we were unable to definitively resolve the relationship between diversification and nodulation. We did, however, succeed in compiling a very large searchable, analysis-ready database of nodulation data for 749 legume genera (98% of Leguminosae genera; LPWG 2017), which, along with associated phylogenetic information, will provide a valuable resource for future work addressing this question and others. For each legume genus, we provide information about the species richness, frequency of nodulation, subfamily association, and topological correspondence with an additional data set of 100 phylogenetic trees curated for database compatibility. We found 386 legume genera were confirmed nodulators (i.e., all species examined for nodulation nodulated), 116 were non-nodulating, four were variable (i.e., containing both confirmed nodulators and confirmed non-nodulators), and 243 had not been examined for nodulation in published studies. Interestingly, data exploration revealed that nodulating legume genera are ~3 × more species-rich than non-nodulating genera, but we did not find evidence that this difference in diversity was due to differences in net diversification rate. Our metadata file describes in more detail the structure of these data that provide a foundational resource for future work as more nodulation data become available, and as greater phylogenetic resolution of this ca. 19,500-species family comes into focus. We release this data set under the Creative Commons 4.0 Attribution-ShareAlike License (https://creativecommons.org/licenses/by-sa/4.0/). The data may be used, distributed, and reproduced with proper citation of this article.

Pattern and Process in the Comparative Study of Convergent Evolution.

Understanding processes that have shaped broad-scale biodiversity patterns is a fundamental goal in evolutionary biology. The development of phylogenetic comparative methods has yielded a tool kit for analyzing contemporary patterns by explicitly modeling processes of change in the past, providing neontologists tools for asking questions previously accessible only for select taxa via the fossil record or laboratory experimentation. The comparative approach, however, differs operationally from alternative approaches to studying convergence in that, for studies of only extant species, convergence must be inferred using evolutionary process models rather than being directly measured. As a result, investigation of evolutionary pattern and process cannot be decoupled in comparative studies of convergence, even though such a decoupling could in theory guard against adaptationist bias. Assumptions about evolutionary process underlying comparative tools can shape the inference of convergent pattern in sometimes profound ways and can color interpretation of such patterns. We discuss these issues and other limitations common to most phylogenetic comparative approaches and suggest ways that they can be avoided in practice. We conclude by promoting a multipronged approach to studying convergence that integrates comparative methods with complementary tests of evolutionary mechanisms and includes ecological and biogeographical perspectives. Carefully employed, the comparative method remains a powerful tool for enriching our understanding of convergence in macroevolution, especially for investigation of why convergence occurs in some settings but not others.

Evolution in a Community Context: On Integrating Ecological Interactions and Macroevolution.

Despite a conceptual understanding that evolution and species interactions are inextricably linked, it remains challenging to study ecological and evolutionary dynamics together over long temporal scales. In this review, we argue that, despite inherent challenges associated with reconstructing historical processes, the interplay of ecology and evolution is central to our understanding of macroevolution and community coexistence, and cannot be safely ignored in community and comparative phylogenetic studies. We highlight new research avenues that foster greater consideration of both ecological and evolutionary dynamics as processes that occur along branches of phylogenetic trees. By promoting new ways forward using this perspective, we hope to inspire further integration that creatively co-utilizes phylogenies and ecological data to study eco-evolutionary dynamics over time and space.

Macroevolution of perfume signalling in orchid bees.

Theory predicts that both stabilising selection and diversifying selection jointly contribute to the evolution of sexual signalling traits by (1) maintaining the integrity of communication signals within species and (2) promoting the diversification of traits among lineages. However, for many important signalling traits, little is known about whether these dynamics translate into predictable macroevolutionary signatures. Here, we test for macroevolutionary patterns consistent with sexual signalling theory in the perfume signals of neotropical orchid bees, a group well studied for their chemical sexual communication. Our results revealed both high species-specificity and elevated rates of evolution in perfume signals compared to nonsignalling traits. Perfume complexity was correlated with the number of congeners in a species' range, suggesting that perfume evolution may be tied to the remarkably high number of orchid bee species coexisting together in some neotropical communities. Finally, sister-pair comparisons were consistent with both rapid divergence at speciation and character displacement upon secondary contact. Together, our results provide new insight into the macroevolution of sexual signalling in insects.

Foliar nectar enhances plant-mite mutualisms: the effect of leaf sugar on the control of powdery mildew by domatia-inhabiting mites.

BACKGROUND AND AIMS: Mite domatia are small structures on the underside of plant leaves that provide homes for predacious or fungivorous mites. In turn, mites inhabiting domatia defend the plant by consuming leaf herbivores and pathogens, which can result in a domatia-mediated, plant-mite defence mutualism. Several recent studies have suggested that plants receive enhanced benefits when they provide a foliar food source, such as sugars secreted from extrafloral nectaries, to mite mutualists alongside mite domatia. However, the effect of foliar sugar on reducing leaf pathogen load via domatia-inhabiting mites has not been directly investigated. METHODS: To fill this gap, the links between foliar sugar addition, domatia-inhabiting mite abundance, and pathogen load were experimentally evaluated in wild grape. Furthermore, because the proposed combined benefits of providing food and housing have been hypothesized to select for the evolutionary correlation of extrafloral nectaries and domatia across plant lineages, a literature survey aimed at determining the overlap of mite domatia and extrafloral nectaries across plant groups was also conducted. KEY RESULTS: It was found that leaves with artificial addition of foliar sugar had 58-80 % more mites than leaves without foliar sugar addition, and that higher mite abundances translated to reduced powdery mildew (Erysiphe necator) loads on leaves. It was found that mite domatia and extrafloral nectaries occur non-randomly in the same clades across Eudicots. Genera with both traits are reported to highlight candidate lineages for future studies. CONCLUSIONS: Together, the results demonstrate that foliar sugar can indeed enhance the efficacy of domatia-mediated plant-mite mutualisms, and suggest that this synergism has the potential to influence the co-distribution of foliar nectar and mite domatia across plants.

Olfactory specialization for perfume collection in male orchid bees.

Insects rely on the olfactory system to detect a vast diversity of airborne molecules in their environment. Highly sensitive olfactory tuning is expected to evolve when detection of a particular chemical with great precision is required in the context of foraging and/or finding mates. Male neotropical orchid bees (Euglossini) collect odoriferous substances from multiple sources, store them in specialized tibial pouches and later expose them at display sites, presumably as mating signals to females. Previous analysis of tibial compounds among sympatric species revealed substantial chemical disparity in chemical composition among lineages with outstanding divergence between closely related species. Here, we tested whether specific perfume phenotypes coevolve with matching olfactory adaptations in male orchid bees to facilitate the location and harvest of species-specific perfume compounds. We conducted electroantennographic (EAG) measurements on males of 15 sympatric species in the genus Euglossa that were stimulated with 18 compounds present in variable proportions in male hind tibiae. Antennal response profiles were species-specific across all 15 species, but there was no conspicuous differentiation between closely related species. Instead, we found that the observed variation in EAG activity follows a Brownian motion model of trait evolution, where the probability of differentiation increases proportionally with lineage divergence time. However, we identified strong antennal responses for some chemicals that are present as major compounds in the perfume of the same species, thus suggesting that sensory specialization has occurred within multiple lineages. This sensory specialization was particularly apparent for semi-volatile molecules ('base note' compounds), thus supporting the idea that such compounds play an important role in chemical signaling of euglossine bees. Overall, our study found no close correspondence between antennal responses and behavioral preferences/tibial contents, but confirms the utility of EAG profiling for discovering certain behaviorally active compounds.

On the study of plant defence and herbivory using comparative approaches: how important are secondary plant compounds.

Species comparisons are a cornerstone of biology and there is a long tradition of using the comparative framework to study the ecology and evolution of plant defensive traits. Early comparative studies led to the hypothesis that plant chemistry plays a central role in plant defence, and the evolution of plant secondary chemistry in response to insect herbivory remains a classic example of coevolution. However, recent comparative work has disagreed with this paradigm, reporting little connection between plant secondary chemicals and herbivory across distantly related plant taxa. One conclusion of this new work is that the importance of secondary chemistry in plant defence may have been generally overstated in earlier research. Here, we attempt to reconcile these contradicting viewpoints on the role of plant chemistry in defence by critically evaluating the use and interpretation of species correlations as a means to study defence-herbivory relationships. We conclude that the notion that plant primary metabolites (e.g. leaf nitrogen content) are the principal determinants of herbivory (or the target of natural selection by herbivores) is not likely to be correct. Despite the inference of recent community-wide studies of herbivory, strong evidence remains for a prime role of secondary compounds in plant defence against herbivores.

Defense mutualisms enhance plant diversification.

The ability of plants to form mutualistic relationships with animal defenders has long been suspected to influence their evolutionary success, both by decreasing extinction risk and by increasing opportunity for speciation through an expanded realized niche. Nonetheless, the hypothesis that defense mutualisms consistently enhance plant diversification across lineages has not been well tested due to a lack of phenotypic and phylogenetic information. Using a global analysis, we show that the >100 vascular plant families in which species have evolved extrafloral nectaries (EFNs), sugar-secreting organs that recruit arthropod mutualists, have twofold higher diversification rates than families that lack species with EFNs. Zooming in on six distantly related plant clades, trait-dependent diversification models confirmed the tendency for lineages with EFNs to display increased rates of diversification. These results were consistent across methodological approaches. Inference using reversible-jump Markov chain Monte Carlo (MCMC) to model the placement and number of rate shifts revealed that high net diversification rates in EFN clades were driven by an increased number of positive rate shifts following EFN evolution compared with sister clades, suggesting that EFNs may be indirect facilitators of diversification. Our replicated analysis indicates that defense mutualisms put lineages on a path toward increased diversification rates within and between clades, and is concordant with the hypothesis that mutualistic interactions with animals can have an impact on deep macroevolutionary patterns and enhance plant diversity.

The phylogenetic distribution of extrafloral nectaries in plants.

BACKGROUND AND AIMS: Understanding the evolutionary patterns of ecologically relevant traits is a central goal in plant biology. However, for most important traits, we lack the comprehensive understanding of their taxonomic distribution needed to evaluate their evolutionary mode and tempo across the tree of life. Here we evaluate the broad phylogenetic patterns of a common plant-defence trait found across vascular plants: extrafloral nectaries (EFNs), plant glands that secrete nectar and are located outside the flower. EFNs typically defend plants indirectly by attracting invertebrate predators who reduce herbivory. METHODS: Records of EFNs published over the last 135 years were compiled. After accounting for changes in taxonomy, phylogenetic comparative methods were used to evaluate patterns of EFN evolution, using a phylogeny of over 55 000 species of vascular plants. Using comparisons of parametric and non-parametric models, the true number of species with EFNs likely to exist beyond the current list was estimated. KEY RESULTS: To date, EFNs have been reported in 3941 species representing 745 genera in 108 families, about 1-2 % of vascular plant species and approx. 21 % of families. They are found in 33 of 65 angiosperm orders. Foliar nectaries are known in four of 36 fern families. Extrafloral nectaries are unknown in early angiosperms, magnoliids and gymnosperms. They occur throughout monocotyledons, yet most EFNs are found within eudicots, with the bulk of species with EFNs being rosids. Phylogenetic analyses strongly support the repeated gain and loss of EFNs across plant clades, especially in more derived dicot families, and suggest that EFNs are found in a minimum of 457 independent lineages. However, model selection methods estimate that the number of unreported cases of EFNs may be as high as the number of species already reported. CONCLUSIONS: EFNs are widespread and evolutionarily labile traits that have repeatedly evolved a remarkable number of times in vascular plants. Our current understanding of the phylogenetic patterns of EFNs makes them powerful candidates for future work exploring the drivers of their evolutionary origins, shifts, and losses.

Phylogenetic and experimental tests of interactions among mutualistic plant defense traits in Viburnum (adoxaceae).

Plant traits that mediate mutualistic interactions are widespread, yet few studies have linked their macroevolutionary patterns with the ecological interactions they mediate. Here we merged phylogenetic and experimental approaches to investigate the evolution of two common mutualistic plant traits, extrafloral nectaries (EFNs) and leaf domatia. By using the flowering plant clade Viburnum, we tested whether macroevolutionary patterns support adaptive hypotheses and conducted field surveys and manipulative experiments to examine whether ecological interactions are concordant with evolutionary predictions. Phylogenetic reconstructions suggested that EFN-bearing species are monophyletic, whereas the evolution of domatia correlated with leaf production strategy (deciduous or evergreen) and climate. Domatia were also more common in the EFN clade, suggesting that the two traits may jointly mediate ecological interactions. This result was further investigated in a common-garden survey, where plants with domatia and EFNs on the leaf blade had more mutualistic mites than plants with other trait combinations, and in manipulative field experiments, where the traits additively increased mutualist abundance. Taken together, our results suggest that mutualistic traits in Viburnum are not ecologically independent, as they work in concert to attract and retain mutualists, and their long-term evolution may be influenced by complex interactions among multiple traits, mutualists, and geography.

Phylogeny, ecology, and the coupling of comparative and experimental approaches.

Recent progress in the development of phylogenetic methods and access to molecular phylogenies has made comparative biology more popular than ever before. However, determining cause and effect in phylogenetic comparative studies is inherently difficult without experimentation and evolutionary replication. Here, we provide a roadmap for linking comparative phylogenetic patterns with ecological experiments to test causal hypotheses across ecological and evolutionary scales. As examples, we consider five cornerstones of ecological and evolutionary research: tests of adaptation, tradeoffs and synergisms among traits, coevolution due to species interactions, trait influences on lineage diversification, and community assembly and composition. Although several scenarios can result in a lack of concordance between historical patterns and contemporary experiments, we argue that the coupling of phylogenetic and experimental methods is an increasingly revealing approach to hypothesis testing in evolutionary ecology.

Toxic cardenolides: chemical ecology and coevolution of specialized plant-herbivore interactions.

Cardenolides are remarkable steroidal toxins that have become model systems, critical in the development of theories for chemical ecology and coevolution. Because cardenolides inhibit the ubiquitous and essential animal enzyme Na⁺/K⁺-ATPase, most insects that feed on cardenolide-containing plants are highly specialized. With a huge diversity of chemical forms, these secondary metabolites are sporadically distributed across 12 botanical families, but dominate the Apocynaceae where they are found in > 30 genera. Studies over the past decade have demonstrated patterns in the distribution of cardenolides among plant organs, including all tissue types, and across broad geographic gradients within and across species. Cardenolide production has a genetic basis and is subject to natural selection by herbivores. In addition, there is strong evidence for phenotypic plasticity, with the biotic and abiotic environment predictably impacting cardenolide production. Mounting evidence indicates a high degree of specificity in herbivore-induced cardenolides in Asclepias. While herbivores of cardenolide-containing plants often sequester the toxins, are aposematic, and possess several physiological adaptations (including target site insensitivity), there is strong evidence that these specialists are nonetheless negatively impacted by cardenolides. While reviewing both the mechanisms and evolutionary ecology of cardenolide-mediated interactions, we advance novel hypotheses and suggest directions for future work.