Carolina Granados Mendoza

Application of the phylogenetic informativeness method to chloroplast markers: a test case of closely related species in tribe Hydrangeeae (Hydrangeaceae).

In evolutionary biology appropriate marker selection for the reconstruction of solid phylogenetic hypotheses is fundamental. One of the most challenging tasks addresses the appropriate choice of genomic regions in studies of closely related species. Robust phylogenetic frameworks are central to studies dealing with questions ranging from evolutionary and conservation biology, biogeography to plant breeding. Phylogenetic informativeness profiles provide a quantitative measure of the phylogenetic signal in markers and therefore a method for locus prioritization. The present work profiles phylogenetic informativeness of mostly non-coding chloroplast regions in an angiosperm lineage of closely related species: the popular ornamental tribe Hydrangeeae (Hydrangeaceae, Cornales, Asterids). A recent phylogenetic study denoted a case of resolution contrast between the two strongly supported clades within tribe Hydrangeeae. We evaluate the phylogenetic signal of 13 highly variable plastid markers for estimating relationships within and among the currently recognized monophyletic groups of this tribe. A selection of combined loci based on their phylogenetic informativeness retrieved more robust phylogenetic hypotheses than simply combining individual markers performing best with respect to resolution, nodal support and accuracy or those presenting the highest number of parsimony informative characters. We propose the rpl32-ndhF intergenic spacer (IGS), trnV-ndhC IGS, trnL-rpl32 IGS, psbT-petB region and ndhA intron as the best candidates for future phylogenetic studies in Hydrangeeae and potentially in other Asterids. We also contrasted the phylogenetic informativeness of coded indels against substitutions concluding that, despite their low phylogenetic informativeness, coded indels provide additional phylogenetic signal that is nearly free of noise. Phylogenetic relationships obtained from our total combined analyses showed improved resolution and nodal support with respect to recently published results.

A genome-scale mining strategy for recovering novel rapidly-evolving nuclear single-copy genes for addressing shallow-scale phylogenetics in Hydrangea

BackgroundIdentifying orthologous molecular markers that potentially resolve relationships at and below species level has been a major challenge in molecular phylogenetics over the past decade. Non-coding regions of nuclear low- or single-copy markers are a vast and promising source of data providing information for shallow-scale phylogenetics. Taking advantage of public transcriptome data from the One Thousand Plant Project (1KP), we developed a genome-scale mining strategy for recovering potentially orthologous single-copy markers to address low-scale phylogenetics. Our marker design targeted the amplification of intron-rich nuclear single-copy regions from genomic DNA. As a case study we used Hydrangea section Cornidia, one of the most recently diverged lineages within Hydrangeaceae (Cornales), for comparing the performance of three of these nuclear markers to other “fast” evolving plastid markers.ResultsOur data mining and filtering process retrieved 73 putative nuclear single-copy genes which are potentially useful for resolving phylogenetic relationships at a range of divergence depths within Cornales. The three assessed nuclear markers showed considerably more phylogenetic signal for shallow evolutionary depths than conventional plastid markers. Phylogenetic signal in plastid markers increased less markedly towards deeper evolutionary divergences. Potential phylogenetic noise introduced by nuclear markers was lower than their respective phylogenetic signal across all evolutionary depths. In contrast, plastid markers showed higher probabilities for introducing phylogenetic noise than signal at the deepest evolutionary divergences within the tribe Hydrangeeae (Hydrangeaceae).ConclusionsWhile nuclear single-copy markers are highly informative for shallow evolutionary depths without introducing phylogenetic noise, plastid markers might be more appropriate for resolving deeper-level divergences such as the backbone relationships of the Hydrangeaceae family and deeper, at which non-coding parts of nuclear markers could potentially introduce noise due to elevated rates of evolution. The herein developed and demonstrated transcriptome based mining strategy has a great potential for the design of novel and highly informative nuclear markers for a range of plant groups and evolutionary scales.

Molecular phylogenetics and new (infra)generic classification to alleviate polyphyly in tribe Hydrangeeae (Cornales: Hydrangeaceae)

Tribe Hydrangeeae of Hydrangeaceae currently contains nine morphologically diverse genera, many of which are well-known garden ornamentals. Previous studies have shown eight of these genera to be phylogenetically nested within Hydrangea, rendering the latter polyphyletic. To clarify the phylogeny of tribe Hydrangeeae, the present study sequenced four chloroplast regions and ITS for an extensive set of taxa, including the type for all nine genera involved. The resulting phylogenetic hypotheses corroborate the polyphyly of Hydrangea. Since polyphyletic taxa are deemed unacceptable by both sides in the ongoing debate concerning the adherence to strict monophyly in biological classifications, a new (infra)generic classification for tribe Hydrangeeae is proposed. In order to create a stable, evolutionary informative classification a broader circumscription of the genus Hydrangea is proposed, to include all eight satellite genera of the tribe. Such treatment is considered highly preferable to an alternative where Hydrangea is to be split into several morphologically potentially unidentifiable genera. To facilitate the acceptance of the new classification proposed here, and in order to create a classification with high information content, the familiar generic names were maintained as section names where possible.

Recalcitrant deep and shallow nodes in Aristolochia (Aristolochiaceae) illuminated using anchored hybrid enrichment

Recalcitrant relationships are characterized by very short internodes that can be found among shallow and deep phylogenetic scales all over the tree of life. Adding large amounts of presumably informative sequences, while decreasing systematic error, has been suggested as a possible approach to increase phylogenetic resolution. The development of enrichment strategies, coupled with next generation sequencing, resulted in a cost-effective way to facilitate the reconstruction of recalcitrant relationships. By applying the anchored hybrid enrichment (AHE) genome partitioning strategy to Aristolochia using an universal angiosperm probe set, we obtained 231-233 out of 517 single or low copy nuclear loci originally contained in the enrichment kit, resulting in a total alignment length of 154,756bp to 160,150bp. Since Aristolochia (Piperales; magnoliids) is distantly related to any angiosperm species whose genome has been used for the plant AHE probe design (Amborella trichopoda being the closest), it serves as a proof of universality for this probe set. Aristolochia comprises approximately 500 species grouped in several clades (OTUs), whose relationships to each other are partially unknown. Previous phylogenetic studies have shown that these lineages branched deep in time and in quick succession, seen as short-deep internodes. Short-shallow internodes are also characteristic of some Aristolochia lineages such as Aristolochia subsection Pentandrae, a clade of presumably recent diversification. This subsection is here included to test the performance of AHE at species level. Filtering and subsampling loci using the phylogenetic informativeness method resolves several recalcitrant phylogenetic relationships within Aristolochia. By assuming different ploidy levels during bioinformatics processing of raw data, first hints are obtained that polyploidization contributed to the evolution of Aristolochia. Phylogenetic results are discussed in the light of current systematics and morphology.

Facilitating wide hybridization in Hydrangea s. l. cultivars: A phylogenetic and marker-assisted breeding approach

Hydrangea s. l., belonging to the up-market segment of ornamental cultivars, currently faces a renaissance in horticulture. Hence, novel molecular-assisted breeding approaches are timely. Wide hybridization, i.e. crosses between distantly related species, has been shown to be problematic. Recent studies have considerably improved our knowledge of the phylogenetic relationships between the ornamental Hydrangea s. l. species. A fully resolved and highly supported phylogenetic tree is currently available, based on an extensive marker selection including 13 highly variable chloroplast markers. This robust phylogenetic framework includes the majority of widely cultivated Hydrangea s. l. species that have been the center of attention in a number of crossing projects. The present study is based on this highly supported phylogenetic hypothesis. Here, we aim to select the best candidates for future successful breeding projects, involving interspecific crosses of both closely and distantly related Hydrangea s. l. lineages. Therefore, we integrated the phylogenetic relatedness of potential parental lines along with genetic distances calculated from a wide plastid marker selection. Direct crosses between two species were found to be successful up to an average genetic distance of 0.01065, while failure could be expected at an average genetic distance of 0.01385 and higher. In order to overcome this genetic distance threshold, we propose Hydrangea arborescens, H. sargentiana, H. integrifolia, and H. seemannii as the best candidates for future bridge-cross projects with currently available fertile hybrids. We expect that our results will allow breeders to overcome long-standing wide crossing difficulties and motivate breeding initiatives of potential economic value.

Phylogeny of the Neotropical sages (Salvia subg. Calosphace; Lamiaceae) and insights into pollinator and area shifts

Salvia subg. Calosphace (Lamiaceae, Lamiales) is a highly diverse clade endemic to the New World. The phylogenetic relationships of Calosphace have been previously investigated using DNA sequences of nuclear ITS region and plastid psbA–trnH intergenic spacer, but the resulting trees lack resolution and support for many clades. The present paper reassesses the phylogenetic relationships of subgenus Calosphace, including a broader taxon sampling, with a special focus on representing previously unsampled sections, and using an additional plastid marker (trnL–trnF region). Our results show increased resolution and overall patterns of support, recovering ten main clades. Within core Calosphace, the most inclusive of these main clades, 17 new subclades were identified. Of the 42 sections for which more than one species was analysed, only 12 are monophyletic. Our biogeographical analysis identified at least twelve migrations to South America from Mexican and Central American lineages, in agreement with previous suggestions of multiple origins of South American Calosphace diversity. This analysis also confirmed a colonization of the Antilles by Andean lineages. The reconstruction of ancestral states of pollination syndromes showed multiple shifts to ornithophily from melittophily and one reversal to the latter.

Bouldering: an alternative strategy to long-vertical climbing in root-climbing hortensias

In the Neotropics, the genus Hydrangea of the popular ornamental hortensia family is represented by climbing species that strongly cling to their support surface by means of adhesive roots closely positioned along specialized anchoring stems. These root-climbing hortensia species belong to the nearly exclusive American Hydrangea section Cornidia and generally are long lianescent climbers that mostly flower and fructify high in the host tree canopy. The Mexican species Hydrangea seemannii, however, encompasses not only long lianescent climbers of large vertical rock walls and coniferous trees, but also short ‘shrub-like’ climbers on small rounded boulders. To investigate growth form plasticity in root-climbing hortensia species, we tested the hypothesis that support variability (e.g. differences in size and shape) promotes plastic responses observable at the mechanical, structural and anatomical level. Stem bending properties, architectural axis categorization, tissue organization and wood density were compared between boulder and long-vertical tree-climbers of H. seemannii. For comparison, the mechanical patterns of a closely related, strictly long-vertical tree-climbing species were investigated. Hydrangea seemannii has fine-tuned morphological, mechanical and anatomical responses to support variability suggesting the presence of two alternative root-climbing strategies that are optimized for their particular environmental conditions. Our results suggest that variation of some stem anatomical traits provides a buffering effect that regulates the mechanical and hydraulic demands of two distinct plant architectures. The adaptive value of observed plastic responses and the importance of considering growth form plasticity in evolutionary and conservation studies are discussed.

Multilocus coalescent species delimitation to evaluate traditionally defined morphotypes in Hydrangea sect. Asperae (Hydrangeaceae)

The number of species recognized in section Asperae of the flowering plant genus Hydrangea differs widely between subsequent revisions. This variation is largely centered around the H. aspera species complex, with numbers of recognized species varying from one to nearly a dozen. Despite indications of molecular variation in this complex, no sequence-based species delimitation methods have been employed to evaluate the primarily morphology-based species boundaries. In the present study, a multi-locus coalescent-based approach to species delimitation is employed in order to identify separate evolutionary lines within H. sect. Asperae, using four chloroplast and four nuclear molecular markers. Eight lineages were recovered within the focal group, of which five correspond with named morphotypes. The other three lineages illustrate types of conflict between molecular species delimitation and traditional morphology-based taxonomy. One molecular lineage comprises two named morphotypes, which possibly diverged recently enough to not have developed sufficient molecular divergence. A second conflict is found in H. strigosa. This morphotype is recovered as a separate lineage when occurring in geographic isolation, but when occurring in sympatry with two other morphotypes (H. aspera and H. robusta), the coalescent species delimitation lumps these taxa into a single putative species.