Stacey D. Smith

The Role of Pollinator Shifts in the Floral Diversification of Iochroma (Solanaceae)

Abstract Differences in floral traits among plant species have often been attributed to adaptation to pollinators. We explored the importance of pollinator shifts in explaining floral divergence among 15 species of Iochroma. We examined four continuously varying floral traits: corolla length, nectar reward, display size, and flower color. Pollinator associations were characterized with a continuously varying measure of pollinator importance (the product of visitation and pollen deposition) for four groups of pollinators: hummingbirds, Hymenoptera, Lepidoptera, and Diptera. A phylogenetic generalized least squares approach was used to estimate correlations between pollinator groups and floral traits across a sample of Bayesian trees using different models of trait evolution. Multivariate analyses were also employed to identify suites of traits associated with each pollinator group. We found that nonphylogenetic models typically fit the data better than phylogenetic models (Brownian motion, Ornstein–Uhlenbeck), and thus results varied little across trees. Our results indicated that species with high nectar reward and large displays are significantly more likely to be pollinated by hummingbirds and less likely to be pollinated by all groups of insects. Corolla length and flower color did not show any consistently significant associations with pollinator groups. For these two traits, we discuss alternative evolutionary forces, including phylogenetic inertia and community-level factors.

Phylogenetic relationships of Malvatheca (Bombacoideae and Malvoideae; Malvaceae sensu lato) as inferred from plastid DNA sequences.

Previous molecular phylogenetic analyses have revealed that elements of the former families Malvaceae sensu stricto and Bombacaceae together form a well-supported clade that has been named Malvatheca. Within Malvatheca, two major lineages have been observed; one, Bombacoideae, corresponds approximately to the palmate-leaved Bombacaceae, and the other, Malvoideae, includes the traditional Malvaceae (the mallows or Eumalvoideae). However, the composition of these two groups and their relationships to other elements of Malvatheca remain a source of uncertainty. Sequence data from two plastid regions, ndhF and trnK/matK, from 34 exemplars of Malvatheca and six outgroups were analyzed. Parsimony, likelihood, and Bayesian analyses of the sequence data provided a well-resolved phylogeny except that relationships among five lineages at the base of Malvatheca are poorly resolved. Nonetheless, a 6-bp insertion in matK suggests that Fremontodendreae is sister to the remainder of Malvatheca. Our results suggest that the Malvoideae originated in the Neotropics and that a mangrove taxon dispersed across the Pacific from South America to Australasia and later radiated out of Australasia to give rise to the ca. 1700 living species of Eumalvoideae. Local clock analyses imply that the plastid genome underwent accelerated molecular evolution coincident with the dispersal out of the Americas and again with the radiation into the three major clades of Eumalvoideae.

Phylogenetics of the florally diverse Andean clade Iochrominae (Solanaceae)

Recent molecular phylogenetic studies of Solanaceae have identified many well-supported clades within the family and have permitted the creation of a phylogenetic system of classification. Here we estimate the phylogeny for Iochrominae, a clade of Physaleae sensu Olmstead et al. (1999), which contains 34 Andean species encompassing an immense diversity of floral forms and colors. Using three nuclear regions, ITS, the second intron of LEAFY, and exons 2 to 9 of the granule-bound starch synthase gene (waxy), we evaluated the monophyly of the traditional genera comprising Iochrominae and assessed the extent of interspecific hybridization within the clade. Only one of the six traditionally recognized genera of Iochrominae was supported as monophyletic. Further, comparison of the individual nuclear data sets revealed two interspecific hybrid taxa and a third possible case. These hybrid taxa occur in the Amotape-Huancabamba zone, a region between the northern and central Andes that has the greatest diversity of Iochroma species and offers frequent opportunities for hybridization in areas of sympatry. We postulate that periodic hybridization events in this area coupled with pollinator-mediated selection and the potential for microallopatry may have acted together to promote diversification in montane Andean taxa, such as Iochrominae.

Plant sex and the evolution of plant defenses against herbivores

Despite the importance of plant–herbivore interactions to the ecology and evolution of terrestrial ecosystems, the evolutionary factors contributing to variation in plant defenses against herbivores remain unresolved. We used a comparative phylogenetic approach to examine a previously untested hypothesis (Recombination-Mating System Hypothesis) that posits that reduced sexual reproduction limits adaptive evolution of plant defenses against arthropod herbivores. To test this hypothesis we focused on the evening primrose family (Onagraceae), which includes both sexual and functionally asexual species. Ancestral state reconstructions on a 5-gene phylogeny of the family revealed between 18 and 21 independent transitions between sexual and asexual reproduction. Based on these analyses, we examined susceptibility to herbivores on 32 plant species representing 15 independent transitions. Generalist caterpillars consumed 32% more leaf tissue, gained 13% greater mass, and experienced 21% higher survival on functionally asexual than on sexual plant species. Survival of a generalist feeding mite was 19% higher on asexual species. In a field experiment, generalist herbivores consumed 64% more leaf tissue on asexual species. By contrast, a specialist beetle fed more on sexual than asexual species, suggesting that a tradeoff exists between the evolution of defense to generalist and specialist herbivores. Measures of putative plant defense traits indicate that both secondary compounds and physical leaf characteristics may mediate this tradeoff. These results support the Recombination-Mating System Hypothesis and suggest that variation in sexual reproduction among plant species may play an important, yet overlooked, role in shaping the macroevolution of plant defenses against arthropod herbivores.

Gene Loss and Parallel Evolution Contribute to Species Difference in Flower Color

Although the importance of regulatory and functional sequence evolution in generating species differences has been studied to some extent, much less is known about the role of other types of genomic changes, such as fluctuation in gene copy number. Here, we apply analyses of gene function and expression of anthocyanin pigment pathway genes, as well as cosegregation analyses in backcross populations, to examine the genetic changes involved in the shift from blue to red flowers in Andean Iochroma (Solanaceae). We demonstrate that deletion of a gene coding for an anthocyanin pathway enzyme was necessary for the transition to red floral pigmentation. The downregulation of a second pathway gene was also necessary for the novel flower color, and this regulatory pattern parallels the genetic change in the two other red-flowered species in the sister family Convolvulaceae in which flower color change has been examined genetically. Finally, we document a shift in enzymatic function at a third locus, but the importance of this change in the transition to red flowers depends on the exact order with which the three changes occurred. This study shows that gene inactivation or loss can be involved in the origin of phenotypic differences between species, thereby restricting the possibility of reversion to the ancestral state. It also demonstrates that parallel evolution of red flowers in three different species occurs via a common developmental/regulatory change but by mutations in different genes.

INFLUENCE OF POLLINATION SPECIALIZATION AND BREEDING SYSTEM ON FLORAL INTEGRATION AND PHENOTYPIC VARIATION IN IPOMOEA

Natural selection should reduce phenotypic variation and increase integration of floral traits involved in placement of pollen grains on stigmas. In this study, we examine the role of pollinators and breeding system on the evolution of floral traits by comparing the patterns of floral phenotypic variances and covariances in 20 Ipomoea species that differ in their level of pollination specialization and pollinator dependence incorporating phylogenetic relatedness. Plants with specialized pollination (i.e., those pollinated by one functional group or by few morphospecies) displayed less phenotypic variation and greater floral integration than generalist plants. Self‐compatible species also displayed greater floral integration than self‐incompatible species. Floral traits involved in pollen placement and pick up showed less variation and greater integration than floral traits involved in pollinator attraction. Analytical models indicate that both breeding system and the number of morphospecies had significant effects on floral integration patterns although only differences in the former were significant after accounting for phylogeny. These results suggest that specialist/self‐compatible plants experience more consistent selection on floral traits than generalist/self‐incompatible plants. Furthermore, pollinators and breeding system promote integration of floral traits involved in pollen placement and pick up rather than integration of the whole flower.

Loss of sexual recombination and segregation is associated with increased diversification in evening primroses.

The loss of sexual recombination and segregation in asexual organisms has been portrayed as an irreversible process that commits asexually reproducing lineages to reduced diversification. We test this hypothesis by estimating rates of speciation, extinction, and transition between sexuality and functional asexuality in the evening primroses. Specifically, we estimate these rates using the recently developed BiSSE (Binary State Speciation and Extinction) phylogenetic comparative method, which employs maximum likelihood and Bayesian techniques. We infer that net diversification rates (speciation minus extinction) in functionally asexual evening primrose lineages are roughly eight times faster than diversification rates in sexual lineages, largely due to higher speciation rates in asexual lineages. We further reject the hypothesis that a loss of recombination and segregation is irreversible because the transition rate from functional asexuality to sexuality is significantly greater than zero and in fact exceeded the reverse rate. These results provide the first empirical evidence in support of the alternative theoretical prediction that asexual populations should instead diversify more rapidly than sexual populations because they are free from the homogenizing effects of sexual recombination and segregation. Although asexual reproduction may often constrain adaptive evolution, our results show that the loss of recombination and segregation need not be an evolutionary dead end in terms of diversification of lineages.

Using phylogenetics to detect pollinator-mediated floral evolution.

The development of comparative phylogenetic methods has provided a powerful toolkit for addressing adaptive hypotheses, and researchers have begun to apply these methods to test the role of pollinators in floral evolution and diversification. One approach is to reconstruct the history of both floral traits and pollination systems to determine if floral trait change is spurred by shifts in pollinators. Looking across multiple shifts, it is also possible to test for significant correlations between floral evolution and pollinators using parsimony, likelihood and Bayesian methods for discrete characters or using statistical comparative methods for continuous characters. Evolutionary shifts in pollinators and floral traits may cause changes in diversification rates, and new methods are available for simultaneously studying character evolution and diversification rates. Relatively few studies have yet used formal comparative methods to elucidate how pollinators affect floral evolution across the phylogeny, and fruitful directions for future applications are discussed.

Competition for hummingbird pollination shapes flower color variation in Andean solanaceae.

One classic explanation for the remarkable diversity of flower colors across angiosperms involves evolutionary shifts among different types of pollinators with different color preferences. However, the pollinator shift model fails to account for the many examples of color variation within clades that share the same pollination system. An alternate explanation is the competition model, which suggests that color divergence evolves in response to interspecific competition for pollinators, as a means to decrease interspecific pollinator movements. This model predicts color overdispersion within communities relative to null assemblages. Here, we combine morphometric analyses, field surveys, and models of pollinator vision with a species‐level phylogeny to test the competition model in the primarily hummingbird‐pollinated clade Iochrominae (Solanaceae). Results show that flower color as perceived by pollinators is significantly overdispersed within sites. This pattern is not simply due to phylogenetic history: phylogenetic community structure does not deviate from random expectations, and flower color lacks phylogenetic signal. Moreover, taxa that occur in sympatry occupy a significantly larger volume of color space than those in allopatry, supporting the hypothesis that competition in sympatry drove the evolution of novel colors. We suggest that competition among close relatives may commonly underlie floral divergence, especially in species‐rich habitats where congeners frequently co‐occur.

A 15-Myr-Old Genetic Bottleneck

Balancing selection preserves variation at the self-incompatibility locus (S-locus) of flowering plants for tens of millions of years, making it possible to detect demographic events that occurred prior to the origin of extant species. In contrast to other Solanaceae examined, SI species in the sister genera Physalis and Witheringia share restricted variation at the S-locus. This restriction is indicative of an ancient bottleneck that occurred in a common ancestor. We sequenced 14 S-alleles from the subtribe Iochrominae, a group that is sister to the clade containing Physalis and Witheringia. At least 6 ancient S-allele lineages are represented among these alleles, demonstrating that the Iochrominae taxa do not share the restriction in S-locus diversity. Therefore, the bottleneck occurred after the divergence of the Iochrominae from the lineage leading to the most recent common ancestor of Physalis and Witheringia. Using cpDNA sequences, 3 fossil dates, and a Bayesian-relaxed molecular clock approach, the crown group of Solanaceae was estimated to be 51 Myr old and the restriction of variation at the S-locus occurred 14.0-18.4 Myr before present. These results confirm the great age of polymorphism at the S-locus and the utility of loci under balancing selection for deep historical inference.