Phylogenomics and the rise of the angiosperms.

Angiosperms are the cornerstone of most terrestrial ecosystems and human livelihoods1,2. A robust understanding of angiosperm evolution is required to explain their rise to ecological dominance. So far, the angiosperm tree of life has been determined primarily by means of analyses of the plastid genome3,4. Many studies have drawn on this foundational work, such as classification and first insights into angiosperm diversification since their Mesozoic origins5-7. However, the limited and biased sampling of both taxa and genomes undermines confidence in the tree and its implications. Here, we build the tree of life for almost 8,000 (about 60%) angiosperm genera using a standardized set of 353 nuclear genes8. This 15-fold increase in genus-level sampling relative to comparable nuclear studies9 provides a critical test of earlier results and brings notable change to key groups, especially in rosids, while substantiating many previously predicted relationships. Scaling this tree to time using 200 fossils, we discovered that early angiosperm evolution was characterized by high gene tree conflict and explosive diversification, giving rise to more than 80% of extant angiosperm orders. Steady diversification ensued through the remaining Mesozoic Era until rates resurged in the Cenozoic Era, concurrent with decreasing global temperatures and tightly linked with gene tree conflict. Taken together, our extensive sampling combined with advanced phylogenomic methods shows the deep history and full complexity in the evolution of a megadiverse clade.

Phylogenomic analyses of Sapindales support new family relationships, rapid Mid-Cretaceous Hothouse diversification, and heterogeneous histories of gene duplication.

Sapindales is an angiosperm order of high economic and ecological value comprising nine families, c. 479 genera, and c. 6570 species. However, family and subfamily relationships in Sapindales remain unclear, making reconstruction of the order's spatio-temporal and morphological evolution difficult. In this study, we used Angiosperms353 target capture data to generate the most densely sampled phylogenetic trees of Sapindales to date, with 448 samples and c. 85% of genera represented. The percentage of paralogous loci and allele divergence was characterized across the phylogeny, which was time-calibrated using 29 rigorously assessed fossil calibrations. All families were supported as monophyletic. Two core family clades subdivide the order, the first comprising Kirkiaceae, Burseraceae, and Anacardiaceae, the second comprising Simaroubaceae, Meliaceae, and Rutaceae. Kirkiaceae is sister to Burseraceae and Anacardiaceae, and, contrary to current understanding, Simaroubaceae is sister to Meliaceae and Rutaceae. Sapindaceae is placed with Nitrariaceae and Biebersteiniaceae as sister to the core Sapindales families, but the relationships between these families remain unclear, likely due to their rapid and ancient diversification. Sapindales families emerged in rapid succession, coincident with the climatic change of the Mid-Cretaceous Hothouse event. Subfamily and tribal relationships within the major families need revision, particularly in Sapindaceae, Rutaceae and Meliaceae. Much of the difficulty in reconstructing relationships at this level may be caused by the prevalence of paralogous loci, particularly in Meliaceae and Rutaceae, that are likely indicative of ancient gene duplication events such as hybridization and polyploidization playing a role in the evolutionary history of these families. This study provides key insights into factors that may affect phylogenetic reconstructions in Sapindales across multiple scales, and provides a state-of-the-art phylogenetic framework for further research.

Target enrichment improves phylogenetic resolution in the genus Zanthoxylum (Rutaceae) and indicates both incomplete lineage sorting and hybridization events.

BACKGROUND AND AIMS: Zanthoxylum is the only pantropical genus within Rutaceae, with a few species native to temperate eastern Asia and North America. Efforts using Sanger sequencing failed to resolve the backbone phylogeny of Zanthoxylum. In this study, we employed target-enrichment high-throughput sequencing to improve resolution. Gene trees were examined for concordance and sectional classifications of Zanthoxylum were evaluated. Off-target reads were investigated to identify putative single-copy markers for bait refinement, and low-copy markers for evidence of putative hybridization events. METHODS: A custom bait set targeting 354 genes, with a median of 321 bp, was designed for Zanthoxylum and applied to 44 Zanthoxylum species and one Tetradium species as the outgroup. Illumina reads were processed via the HybPhyloMaker pipeline. Phylogenetic inferences were conducted using coalescent and maximum likelihood methods based on concatenated datasets. Concordance was assessed using quartet sampling. Additional phylogenetic analyses were performed on putative single and low-copy genes extracted from off-target reads. KEY RESULTS: Four major clades are supported within Zanthoxylum: the African clade, the Z. asiaticum clade, the Asian-Pacific-Australian clade and the American-eastern Asian clade. While overall support has improved, regions of conflict are similar to those previously observed. Gene tree discordances indicate a hybridization event in the ancestor of the Hawaiian lineage, and incomplete lineage sorting in the American backbone. Off-target putative single-copy genes largely confirm on-target results, and putative low-copy genes provide additional evidence for hybridization in the Hawaiian lineage. Only two of the five sections of Zanthoxylum are resolved as monophyletic. CONCLUSIONS: Target enrichment is suitable for assessing phylogenetic relationships in Zanthoxylum. Our phylogenetic analyses reveal that current sectional classifications need revision. Quartet tree concordance indicates several instances of reticulate evolution. Off-target reads are proven useful to identify additional phylogenetically informative regions for bait refinement or gene tree based approaches.

Characterization of the Complete Chloroplast Genome Sequences of Four Zanthoxylum L. Species (Sapindales: Rutaceae) from the Caribbean, Madagascar, the Mascarene Islands, and the South Pacific.

Zanthoxylum is a genus of woody plants in the Rutaceae family distributed pantropically, with some species extending to temperate regions in East Asia and North America. Here, we present the complete chloroplast genome sequences of four species, two of them critically endangered, endemic to tropical islands.

New Acetophenones and Chromenes from the Leaves of Melicope barbigera A. Gray.

The dichloromethane extract from leaves of Melicope barbigera (Rutaceae), endemic to the Hawaiian island of Kaua'i, yielded four new and three previously known acetophenones and 2H-chromenes, all found for the first time in M. barbigera. The structures of the new compounds obtained from the dichloromethane extract after purification by chromatographic methods were unambiguously elucidated by spectroscopic analyses including 1D/2D NMR spectroscopy and HRESIMS. The absolute configuration was determined by modified Mosher's method. Compounds 2, 4 and the mixture of 6 and 7 exhibited moderate cytotoxic activities against the human ovarian cancer cell line A2780 with IC50 values of 30.0 and 75.7 µM for 2 and 4, respectively, in a nuclear shrinkage cytotoxicity assay.

New Guinea has the world's richest island flora.

New Guinea is the world's largest tropical island and has fascinated naturalists for centuries1,2. Home to some of the best-preserved ecosystems on the planet3 and to intact ecological gradients-from mangroves to tropical alpine grasslands-that are unmatched in the Asia-Pacific region4,5, it is a globally recognized centre of biological and cultural diversity6,7. So far, however, there has been no attempt to critically catalogue the entire vascular plant diversity of New Guinea. Here we present the first, to our knowledge, expert-verified checklist of the vascular plants of mainland New Guinea and surrounding islands. Our publicly available checklist includes 13,634 species (68% endemic), 1,742 genera and 264 families-suggesting that New Guinea is the most floristically diverse island in the world. Expert knowledge is essential for building checklists in the digital era: reliance on online taxonomic resources alone would have inflated species counts by 22%. Species discovery shows no sign of levelling off, and we discuss steps to accelerate botanical research in the 'Last Unknown'8.

Phylogeny of Hawaiian Melicope (Rutaceae): RAD-seq Resolves Species Relationships and Reveals Ancient Introgression.

Hawaiian Melicope are one of the major adaptive radiations of the Hawaiian Islands comprising 54 endemic species. The lineage is monophyletic with an estimated crown age predating the rise of the current high islands. Phylogenetic inference based on Sanger sequencing has not been sufficient to resolve species or deeper level relationships. Here, we apply restriction site-associated DNA sequencing (RAD-seq) to the lineage to infer phylogenetic relationships. We employ Quartet Sampling to assess information content and statistical support, and to quantify discordance as well as partitioned ABBA-BABA tests to uncover evidence of introgression. Our new results drastically improved resolution of relationships within Hawaiian Melicope. The lineage is divided into five fully supported main clades, two of which correspond to morphologically circumscribed infrageneric groups. We provide evidence for both ancestral and current hybridization events. We confirm the necessity for a taxonomic revision of the Melicope section Pelea, as well as a re-evaluation of several species complexes by combining genomic and morphological data.

Phylogeny and biogeography of the pantropical genus Zanthoxylum and its closest relatives in the proto-Rutaceae group (Rutaceae).

Zanthoxylum L. (prickly ash) is the only genus in the Citrus L. family (Rutaceae) with a pantropical distribution. We present the first detailed phylogenetic and biogeographic study of the genus and its close relatives in the proto-Rutaceae group. Our phylogenetic analyses based on two plastid and two nuclear markers show that the genus Toddalia Juss. is nested within Zanthoxylum, that earlier generic and intrageneric classifications need revision, and that the homochlamydeous flowers of the temperate species of Zanthoxylum are the result of a reduction from heterochlamydeous flowers. The biogeographic analyses reveal a Eurasian origin of Zanthoxylum in the Paleocene or Eocene with successive intercontinental or long-range migrations. Zanthoxylum likely crossed the North Atlantic Land Bridges to colonize the Americas in the Eocene, and migrated back to the Old World probably via the Bering Land Bridge in the Oligocene or Miocene. Zanthoxylum also colonized several Pacific Islands and the Hawaiian clade shows phylogenetic incongruence between the plastid and nuclear datasets, suggesting hybridization. The Hawaiian species are one of the rare examples of endemic Hawaiian lineages that are older than the current main islands.

Publisher Correction: A fossil species of the enigmatic early polypod fern genus Cystodium (Cystodiaceae) in Cretaceous amber from Myanmar.

A correction to this article has been published and is linked from the HTML and PDF versions of this paper. The error has been fixed in the paper.

Publisher Correction: A fossil species of the enigmatic early polypod fern genus Cystodium (Cystodiaceae) in Cretaceous amber from Myanmar.

A correction to this article has been published and is linked from the HTML version of this paper. The error has been fixed in the paper.

A fossil species of the enigmatic early polypod fern genus Cystodium (Cystodiaceae) in Cretaceous amber from Myanmar.

The monospecific fern genus Cystodium (Cystodiaceae; Polypodiales) occurs exclusively in the tropical forests of the Malay Archipelago, the Admiralty Islands, the Louisiade Archipelago, and the Solomon Islands. Divergence time estimates suggest that the genus originated in the Mesozoic; however, fossil evidence to validate this suggestion has been lacking. Amber from Myanmar (Burmese amber) is an important source of new information on the diversity of vascular cryptogams in the Cretaceous. This paper describes the fossil taxon Cystodium sorbifolioides nov. sp. based on a fragment of a fertile leaf preserved in Burmese amber that represents the first fossil evidence of the family Cystodiaceae. Cystodium sorbifolioides is used to obtain a minimum age estimate for the Cystodiaceae and the closely related, monogeneric Lonchitidaceae and Lindsaeaceae. The fossil strengthens the hypothesis that the forest ecosystems of Malesia and Melanesia represent refugia for many tropical plant lineages that originated in the Cretaceous.

Melicope stonei, section Pelea (Rutaceae), a new species from Kaua'i, Hawaiian Islands: with notes on its distribution, ecology, conservation status, and phylogenetic placement.

Melicope stonei K.R. Wood, Appelhans & W.L. Wagner (section Pelea, Rutaceae), a new endemic tree species from Kaua'i, Hawaiian Islands, is described and illustrated with notes on its distribution, ecology, conservation status, and phylogenetic placement. The new species differs from its Hawaiian congeners by its unique combination of distinct carpels and ramiflorous inflorescences arising on stems below the leaves; plants monoecious; leaf blades (5-)8-30 × (4-)6-11 cm, with abaxial surface densely tomentose, especially along midribs; and very long petioles of up to 9 cm. Since its discovery in 1988, 94 individuals have been documented and are confined to a 1.5 km2 region of unique high canopy mesic forest. Melicope stonei represents a new Critically Endangered (CR) single island endemic species on Kaua'i.

Analysis of whole chloroplast genomes from the genera of the Clauseneae, the curry tribe (Rutaceae, Citrus family).

The Clauseneae (Aurantioideae, Rutaceae) is a tribe in the Citrus family that, although economically important as it contains the culinary and medicinally-useful curry tree (Bergera koenigii), has been relatively understudied. Due to the recent significant taxonomic changes made to this tribe, a closer inspection of the genetic relationships among its genera has been warranted. Whole genome skimming was used to generate chloroplast genomes from six species, representing each of the four genera (Bergera, Clausena, Glycosmis, Micromelum) in the Clauseneae tribe plus one closely related outgroup (Merrillia), using the published plastome sequence of Citrus sinensis as a reference. Phylogenetically informative character (PIC) data were analyzed using a genome alignment of the seven species, and variability frequency among the species was recorded for each coding and non-coding region, with the regions of highest variability identified for future phylogenetic studies. Non-coding regions exhibited a higher percentage of variable characters as expected, and the phylogenetic markers ycf1, matK, rpoC2, ndhF, trnS-trnG spacer, and trnH-psbA spacer proved to be among the most variable regions. Other markers that are frequently used in phylogenetic studies, e.g. rps16, atpB-rbcL, rps4-trnT, and trnL-trnF, proved to be far less variable. Phylogenetic analyses of the aligned sequences were conducted using Bayesian inference (MrBayes) and Maximum Likelihood (RAxML), yielding highly supported divisions among the four genera.

Reduction of the Hawaiian genus Platydesma into Melicope section Pelea (Rutaceae) and notes on the monophyly of the section.

Platydesma, an endemic genus to the Hawaiian Islands containing four species, has long been considered of obscure origin. Recent molecular phylogenetic studies have unequivocally placed Platydesma within the widespread genus Melicope as sister to the rest of the Hawaiian species of Melicope. This makes submerging Platydesma into Melicope necessary. We make the necessary new combinations: Melicope cornuta (Hillebr.) Appelhans, K.R. Wood & W.L. Wagner, M. cornuta var. decurrens (B.C.Stone) Appelhans, K.R. Wood & W.L. Wagner, M. remyi (Sherff) Appelhans, K.R. Wood & W.L. Wagner, and M. rostrata (Hillebr.) Appelhans, K.R. Wood & W.L. Wagner. An additional species that has been recognized within Platydesma should now be recognized under its original name M. spathulata A. Gray. All Hawaiian species belong to Melicope section Pelea. Our molecular phylogenetic studies also showed that in addition to merging Platydesma into section Pelea, five species described from New Caledonia need to be excluded from the section in order to achieve monophyly of section Pelea.

Melicope oppenheimeri, section Pelea (Rutaceae), a new species from West Maui, Hawaiian Islands: with notes on its ecology, conservation, and phylogenetic placement.

Melicope oppenheimeri K.R. Wood, Appelhans & W.L. Wagner (section Pelea (A. Gray) Hook. f., Rutaceae), a rare endemic tree from West Maui, Hawaiian Islands, is described and illustrated with notes on its ecology, conservation, and phylogenetic placement. The new species differs from Hawaiian congeners by its carpels basally connate 1/5, narrowed into a strongly reflexed beak 10-15 mm long. It also differs in a combination of leaves with 7-10 pair of secondary veins; cymes to 3 cm long; peduncles 5-6.5 mm long; flowers perfect; capsules 4-9 × 40-52 mm; and a densely appressed short-sericeous ovary. Melicope oppenheimeri is known only from an isolated cliff-base plateau in upper Waihe'e Valley, West Maui. Its discovery brings the number of recognized Melicope J.R. Forst. & G. Forst. species in the Hawaiian Islands to 49. A table is included indicating the conservation status of Hawaiian Melicope and Platydesma H. Mann., which is nested within Melicope sect. Pelea. Melicope oppenheimeri falls into the IUCN Critically Endangered (CR) Red List category.

On the identity of Blanco's Cissus frutescens and its correct name in Melicope (Rutaceae) with neotypification of Cissus arborea Blanco.

The names Cissus frutescens and Cissus arborea have a long history of confusion. Cissus frutescens Blanco belongs to the genus Melicope (Rutaceae) and we herein correct a nomenclatural mistake made by T.G. Hartley in the revision of Melicope. The name Melicope confusa (Merr.) P.S. Liu was accepted for this taxon by Hartley. However, Cissus frutescens Blanco represents the earliest name for this entity and a new combination, Melicope frutescens (Blanco) Appelhans & J.Wen is herein proposed. Neotypification of Cissus arborea Blanco is also provided.

Phylogeny of Acronychia (Rutaceae) and First Insights into Its Historical Biogeography and the Evolution of Fruit Characters.

BACKGROUND: The genus Acronychia (Citrus family, Rutaceae) contains 49 species of trees and shrubs that are found mainly in rain forest. The genus has a large distributional range from mainland southern Asia to Australia and New Caledonia, but most species are endemic to either New Guinea or Australia. This study aimed to provide the first detailed molecular phylogeny of Acronychia and use it to test the taxonomic value of fruit morphological characters, and infer the historical biogeography of the genus. METHODOLOGY: Phylogenetic analyses (Bayesian Inference, Maximum Likelihood) were undertaken on nucleotide sequence data from two plastid (psbA-trnH, trnL-trnF) and three nuclear markers (ETS, ITS, NIAi3) from 29 Acronychia species (59% of the genus) and representatives of related genera. RESULTS AND CONCLUSIONS: The results indicate that the South-East Asian genus Maclurodendron is nested phylogenetically within Acronychia and must be synonymized to render Acronychia monophyletic. Fruit morphological characters have been used previously to infer relationships within Acronychia and our analyses show that these characters are informative for some subclades but are homoplasious for the group as a whole. Apocarpous fruits are the ancestral state in Acronychia and subapocarpous and fully syncarpous fruits are derived. The unisexual flowers of Maclurodendron are derived from bisexual flowers, which are found in all species of Acronychia as well as its relatives. Acronychia probably first evolved on Australia with range expansion to New Guinea via stepping-stone dispersal or direct land connections within the Sahul Shelf, followed by two independent dispersals to areas west of New Guinea. Most species of Acronychia occur in either Australia or New Guinea, but no species occurs in both regions. This is surprising given the close proximity of the landmasses, but might be explained by ecological factors.

Melicope balgooyi Appelhans, W.L. Wagner & K.R. Wood, a new species and new record in Melicope section Melicope (Rutaceae) for the Austral Islands.

Melicope balgooyi, a new species of Melicope (Rutaceae) is described. It is known only from the Austral Islands in the South Pacific (French Polynesia). However, it is not closely related to the other two species previously known from the Austral Islands, which are part of Melicope section Vitiflorae. The new species belongs to Melicope section Melicope and is most closely related to species from New Zealand, the Kermadec Islands, and the Society Islands. The new species has alternate to sub-opposite leaves, which is a very rare arrangement in Melicope and has only been described for two other species of the genus so far.

A molecular phylogeny of Acronychia, Euodia, Melicope and relatives (Rutaceae) reveals polyphyletic genera and key innovations for species richness.

We present the first detailed phylogenetic study of the genus Melicope, the largest genus of the Citrus family (Rutaceae). The phylogenetic analysis sampled about 50% of the 235 accepted species of Melicope as well as representatives of 26 related genera, most notably Acronychia and Euodia. The results based on five plastid and nuclear markers have revealed that Acronychia, Euodia and Melicope are each not monophyletic in their current circumscriptions and that several small genera mainly from Australia and New Caledonia need to be merged with one of the three genera to ensure monophyly at the generic level. The phylogenetic position of the drupaceous Acronychia in relation to Melicope, which has capsular or follicular fruits, remains unclear and Acronychia might be a separate genus or a part of Melicope. The seed coats of Melicope, Acronychia and related genera show adaptations to bird-dispersal, which might be regarded as key innovations for species radiations. Euodia and its relatives, which lack these adaptations, include only about 20 species while the Melicope-Acronychia group consists of about 340 species. The drupaceous genera Comptonella, Dutaillyea, Picrella and Sarcomelicope are nested within Melicope and need to be merged with Melicope. The expanded genus is a prime example of the artificial classification system of Engler, who defined Rutaceous subfamilies mainly based on gynoecial and fruit characters.

The phylogeny of yellow fever virus 17D vaccines.

In recent years the safety of the yellow fever live vaccine 17D came under scrutiny. The focus was on serious adverse events after vaccinations that resemble a wild type infection with yellow fever and whose reasons are still not known. Also the exact mechanism of attenuation of the vaccine remains unknown to this day. In this context, the standards of safety and surveillance in vaccine production and administration have been discussed. Therein embodied was the demand for improved documentation of the derivation of the seed virus used for yellow fever vaccine production. So far, there was just a historical genealogy available that is based on source area and passage level. However, there is a need for a documentation based on molecular information to get better insights into the mechanisms of pathology. In this work we sequenced the whole genome of different passages of the YFV-17D strain used by Crucell Switzerland AG for vaccine production. Using all other publically available 17D full genome sequences we compared the sequence variance of all vaccine strains and oppose a phylogenetic tree based on full genome sequences to the historical genealogy.