A phylogenetic analysis of the Orchidaceae: evidence from rbcL nucleotide

Cladistic parsimony analyses of rbcL nucleotide sequence data from 171 taxa representing nearly all tribes and subtribes of Orchidaceae are presented here. These analyses divide the family into five primary monophyletic clades: apostasioid, cypripedioid, vanilloid, orchidoid, and epidendroid orchids, arranged in that order. These clades, with the exception of the vanilloids, essentially correspond to currently recognized subfamilies. A distinct subfamily, based upon tribe Vanilleae, is supported for Vanilla and its allies. The general tree topology is, for the most part, congruent with previously published hypotheses of intrafamilial relationships; however, there is no evidence supporting the previously recognized subfamilies Spiranthoideae, Neottioideae, or Vandoideae. Subfamily Spiranthoideae is embedded within a single clade containing members of Orchidoideae and sister to tribe Diurideae. Genera representing tribe Tropideae are placed within the epidendroid clade. Most traditional subtribal units are supported within each clade, but few tribes, as currently circumscribed, are monophyletic. Although powerful in assessing monophyly of clades within the family, in this case rbcL fails to provide strong support for the interrelationships of the subfamilies (i.e., along the spine of the tree). The cladograms presented here should serve as a standard to which future morphological and molecular studies can be compared.     

Phylogenetic relationships in Epidendroideae (Orchidaceae), one of the great flowering plant radiations: progressive specialization and diversification

Background and Aims The largest subfamily of orchids, Epidendroideae, represents one of the most significant diversifications among flowering plants in terms of pollination strategy, vegetative adaptation and number of species. Although many groups in the subfamily have been resolved, significant relationships in the tree remain unclear, limiting conclusions about diversification and creating uncertainty in the classification. This study brings together DNA sequences from nuclear, plastid and mitochrondrial genomes in order to clarify relationships, to test associations of key characters with diversification and to improve the classification.Methods Sequences from seven loci were concatenated in a supermatrix analysis for 312 genera representing most of epidendroid diversity. Maximum-likelihood and parsimony analyses were performed on this matrix and on subsets of the data to generate trees and to investigate the effect of missing values. Statistical character-associated diversification analyses were performed.Key Results Likelihood and parsimony analyses yielded highly resolved trees that are in strong agreement and show significant support for many key clades. Many previously proposed relationships among tribes and subtribes are supported, and some new relationships are revealed. Analyses of subsets of the data suggest that the relatively high number of missing data for the full analysis is not problematic. Diversification analyses show that epiphytism is most strongly associated with diversification among epidendroids, followed by expansion into the New World and anther characters that are involved with pollinator specificity, namely early anther inflexion, cellular pollinium stalks and the superposed pollinium arrangement.Conclusions All tested characters show significant association with speciation in Epidendroideae, suggesting that no single character accounts for the success of this group. Rather, it appears that a succession of key features appeared that have contributed to diversification, sometimes in parallel.     

Phylogeny of Malpighiaceae: evidence from chloroplast ndhF and trnl-F nucleotide sequences

The Malpighiaceae are a family of ～1250 species of predominantly New World tropical flowering plants. Infrafamilial classification has long been based on fruit characters. Phylogenetic analyses of chloroplast DNA nucleotide sequences were analyzed to help resolve the phylogeny of Malpighiaceae. A total of 79 species, representing 58 of the 65 currently recognized genera, were studied. The 3' region of the gene ndhF was sequenced for 77 species and the noncoding intergenic spacer region trnL-F was sequenced for 65 species; both sequences were obtained for the outgroup, Humiria (Humiriaceae). Phylogenetic relationships inferred from these data sets are largely congruent with one another and with results from combined analyses. The family is divided into two major clades, recognized here as the subfamilies Byrsonimoideae (New World only) and Malpighioideae (New World and Old World). Niedenzu's tribes are all polyphyletic, suggesting extensive convergence on similar fruit types; only de Jussieu's tribe Gaudichaudieae and Anderson's tribes Acmanthereae and Galphimieae are monophyletic. Fleshy fruits evolved three times in the family and bristly fruits at least three times. Among the wing-fruited vines, which constitute more than half the diversity in the family, genera with dorsal-winged samaras are fairly well resolved, while the resolution of taxa with lateral-winged samaras is poor. The trees suggest a shift from radially symmetrical pollen arrangement to globally symmetrical pollen at the base of one of the clades within the Malpighioideae. The Old World taxa fall into at least six and as many as nine clades.     

An updated classification of Orchidaceae

Since the last classification of Orchidaceae in 2003, there has been major progress in the determination of relationships, and we present here a revised classification including a list of all 736 currently recognized genera. A number of generic changes have occurred in Orchideae (Orchidoideae), but the majority of changes have occurred in Epidendroideae. In the latter, almost all of the problematic placements recognized in the previous classification 11 years ago have now been resolved. In Epidendroideae, we have recognized three new tribes (relative to the last classification): Thaieae (monogeneric) for Thaia, which was previously considered to be the only taxon incertae sedis; Xerorchideae (monogeneric) for Xerorchis; and Wullschlaegelieae for achlorophyllous Wullschlaegelia, which had tentatively been placed in Calypsoeae. Another genus, Devogelia, takes the place of Thaia as incertae sedis in Epidendroideae. Gastrodieae are clearly placed among the tribes in the neottioid grade, with Neottieae sister to the remainder of Epidendroideae. Arethuseae are sister to the rest of the higher Epidendroideae, which is unsurprising given their mostly soft pollinia. Tribal relationships within Epidendroideae have been much clarified by analyses of multiple plastid DNA regions and the low‐copy nuclear gene Xdh. Four major clades within the remainder of Epidendroideae are recognized: Vandeae/Podochileae/Collabieae, Cymbidieae, Malaxideae and Epidendreae, the last now including Calypsoinae (previously recognized as a tribe on its own) and Agrostophyllinae s.s. Agrostophyllinae and Collabiinae were unplaced subtribes in the 2003 classification. The former are now split between two subtribes, Agrostophyllinae s.s. and Adrorhizinae, the first now included in Epidendreae and the second in Vandeae. Collabiinae, also probably related to Vandeae, are now elevated to a tribe along with Podochileae. Malaxis and relatives are placed in Malaxidinae and included with Dendrobiinae in Malaxideae. The increased resolution and content of larger clades, recognized here as tribes, do not support the ‘phylads’ in Epidendroideae proposed 22 years ago by Dressler. © 2014 The Linnean Society of London, Botanical Journal of the Linnean Society, 2015, 177 , 151–174.     

Molecular phylogenetics of Maxillaria and related genera (Orchidaceae: Cymbidieae) based on combined molecular data sets

The orchid genus Maxillaria is one of the largest and most common of neotropical orchid genera, but its current generic boundaries and relationships have long been regarded as artificial. Phylogenetic relationships within subtribe Maxillariinae sensu Dressler (1993) with emphasis on Maxillaria s.l. were inferred using parsimony analyses of individual and combined DNA sequence data. We analyzed a combined matrix of nrITS DNA, the plastid matK gene and flanking trnK intron, and the plastid atpB-rbcL intergenic spacer for 619 individuals representing ca. 354 species. The plastid rpoC1 gene (ca. 2600 bp) was sequenced for 84 selected species and combined in a more limited analysis with the other data sets to provide greater resolution. In a well-resolved, supported consensus, most clades were present in more than one individual analysis. All the currently recognized minor genera of "core" Maxillariinae (Anthosiphon, Chrysocycnis, Cryptocentrum, Cyrtidiorchis, Mormolyca, Pityphyllum, and Trigonidium) are embedded within a polyphyletic Maxillaria s.l. Our results support the recognition of a more restricted Maxillaria, of some previously published segregate genera (Brasiliorchis, Camaridium, Christensonella, Heterotaxis, Ornithidium, Sauvetrea), and of several novel clades at the generic level. These revised monophyletic generic concepts should minimize further nomenclatural changes, encourage monographic studies, and facilitate more focused analyses of character evolution within Maxillariinae.     

Rangewide analysis of fungal associations in the fully mycoheterotrophic Corallorhiza striata complex (Orchidaceae) reveals extreme specificity on ectomycorrhizal Tomentella (Thelephoraceae) across North America

Fully mycoheterotrophic plants offer a fascinating system for studying phylogenetic associations and dynamics of symbiotic specificity between hosts and parasites. These plants frequently parasitize mutualistic mycorrhizal symbioses between fungi and trees. Corallorhiza striata is a fully mycoheterotrophic, North American orchid distributed from Mexico to Canada, but the full extent of its fungal associations and specificity is unknown. Plastid DNA (orchids) and ITS (fungi) were sequenced for 107 individuals from 42 populations across North America to identify C. striata mycobionts and test hypotheses on fungal host specificity. Four largely allopatric orchid plastid clades were recovered, and all fungal sequences were most similar to ectomycorrhizal Tomentella (Thelephoraceae), nearly all to T. fuscocinerea. Orchid-fungal gene trees were incongruent but nonindependent; orchid clades associated with divergent sets of fungi, with a clade of Californian orchids subspecialized toward a narrow Tomentella fuscocinerea clade. Both geography and orchid clades were important determinants of fungal association, following a geographic mosaic model of specificity on Tomentella fungi. These findings corroborate patterns described in other fully mycoheterotrophic orchids and monotropes, represent one of the most extensive plant-fungal genetic investigations of fully mycoheterotrophic plants, and have conservation implications for the >400 plant species engaging in this trophic strategy worldwide.     

Molecular phylogenetics of Prescottiinae s.l. and their close allies (Orchidaceae, Cranichideae) inferred from plastid and nuclear ribosomal DNA sequences

The Andes are a cradle of orchid evolution, but most phylogenetic studies of Orchidaceae in this biodiversity hotspot have dealt with epiphytic epidendroid lineages. Here we present a study on neotropical, terrestrial, orchidoid taxa of Prescottiinae s.l. (8 genera, ～100 species), which are adapted to some of the highest elevation habitats on earth that support orchids. They are currently included within an expanded concept of Cranichidinae in the tribe Cranichideae, but DNA sequence data show that neither Prescottiinae s.l. nor Cranichidinae s.s. are monophyletic. Prescottiinae s.l. consist of two strongly supported lineages: the Altensteinia and Prescottia clades, which have closer affinities to Spiranthinae than to Cranichidinae. The Prescottia clade comprises two well-supported subclades, one including most sampled species of Prescottia and a second one with Pseudocranichis thysanochila sister to Prescottia tubulosa. As a group, they are sister to Spiranthinae. Sister to this pair is the Altensteinia clade comprised of six genera, whose intergeneric relationships are well resolved. Finally, Cranichidinae s.s. is sister to all three of these clades. Morphological and ecological features distinguishing the major groups are discussed, as are potential synapomorphies to define them. The reconstructed phylogeny indicates that the classification of Cranichideae needs to be reexamined.     

Chloroplast DNA restriction site variation in theVernonieae (Asteraceae), an initial appraisal of the relationship of New and Old World taxa and the monophyly ofVernonia

Chloroplast DNA restriction site variation was examined for 35 taxa in theVernonieae and four outgroup tribes, using 17 restriction enzymes mapped for ca. 900 restriction sites per species; 139 mutations were found to be phylogenetically informative. Phylogenetic trees were constructed using Wagner and weighted parsimony, and evaluated by bootstrap and decay analyses. Relationships of Old and New World taxa indicate complex geographical relationships; there was no clear geographic separation by hemisphere. The relationships between Old and New World Vernonias found here support prior morphological analyses. The sister group to all New and most Old World taxa was composed of a small group of Old World species including yellow-flowered, trinervate-leaved species previously postulated to be basal in the tribe. The majority of both New and Old World taxa are derived from a lineage beginning with the monotypic genusStokesia, an endemic of the southeastern United States. The genusVernonia was also found to be paraphyletic within both the New and Old World. Available data do not support either the separation ofVernonia or the tribeVernonieae into geographically distinct lineages. The pattern of relationships within theVernonieae for taxa from North America, Asia, Africa, Central and South America is most similar to that of several other groups of both plants and animals with a boreotropical origin, rather than an origin in Gondwanaland. Such a pattern of distribution suggests more ancient vicariant events than are routinely postulated for theAsteraceae.     

北京松山国家级自然保护区兰科植物多样性及其保护评价

选取北京松山国家级自然保护区开展兰科植物多样性调查和分析,评估其保护现状。松山共有兰科植物12属18种,其中北方盔花兰为北京新记录种。在生活型方面,以地生兰为主,计14种（77．78％）,腐生兰4种（22．22％）;在分布区类型方面,属的区系成分以北温带分布为主（6属,5000％）,兼有旧世界温带分布（3属,25．00％）、世界分布（2属,16．67％）和旧世界热带分布（1属,8．33％）;种的区系成分以东亚分布为主（9种,50．00％）,兼有旧世界温带分布（5种,27．78％）和北温带分布（4种,22．22％）。松山兰科植物集中分布于北沟和小海坨山。通过综合分析松山兰科植物保护现状,提出生境评估、种群动态监测、建立迁地保护资源圃和加大宣传等都是切实加强兰科植物保护工作的可行措施。     

Tribal delimitation and phylogenetic relationships of Loteae and Coronilleae (Faboideae: Fabaceae) with special reference to Lotus: evidence from nuclear ribosomal ITS sequences

The temperate herbaceous tribes Loteae and Coronilleae have traditionally been regarded as taxonomically distinct entities. More recent morphological assessments, however, have challenged this view and suggest combining the two tribes under Loteae. Two key features used to distinguish the Coronilleae from Loteae include jointed fruits and branched root nodules. We evaluate the taxonomic utility of these characters using information derived from phylogenetic analyses of the internal transcribed spacers ITS1 + 2, and the intervening 5.8S region of nuclear ribosomal DNA. Results from this study show that neither the Loteae nor Coronilleae form individual monophyletic groups, and that key fruit and root nodule characters used to distinguish the Coronilleae are homoplastic. Given these data, we support the recognition of a single tribe, Loteae. We also find that Lotus, the largest and most morphologically complex genus in either tribe, is not monophyletic. Rather, it consists of two geographically distinct lineages, Old and New World, each of which are more closely related to other Loteae genera: Old World Lotus are more closely related to Old World Anthyllis, while New World Lotus show closer affinities to Old World Coronilla. These data also have important implications for the biogeography of New World Lotus: equally most parsimonious reconstructions suggest a complex scenario of intercontinental dispersals that involve not only Old World Lotus but Coronilla as well.     

Mixotrophy of Platanthera minor, an orchid associated with ectomycorrhiza-forming Ceratobasidiaceae fungi

? We investigated the fungal symbionts and carbon nutrition of a Japanese forest photosynthetic orchid, Platanthera minor, whose ecology suggests a mixotrophic syndrome, that is, a mycorrhizal association with ectomycorrhiza (ECM)-forming fungi and partial exploitation of fungal carbon. ? We performed molecular identification of symbionts by PCR amplifications of the fungal ribosomal DNA on hyphal coils extracted from P. minor roots. We tested for a (13)C and (15)N enrichment characteristic of mixotrophic plants. We also tested the ectomycorrhizal abilities of orchid symbionts using a new protocol of direct inoculation of hyphal coils onto roots of Pinus densiflora seedlings. ? In phylogenetic analyses, most isolated fungi were close to ECM-forming Ceratobasidiaceae clades previously detected from a few fully heterotrophic orchids or environmental ectomycorrhiza surveys. The direct inoculation of fungal coils of these fungi resulted in ectomycorrhiza formation on P. densiflora seedlings. Stable isotope analyses indicated mixotrophic nutrition of P. minor, with fungal carbon contributing from 50% to 65%. ? This is the first evidence of photosynthetic orchids associated with ectomycorrhizal Ceratobasidiaceae taxa, confirming the evolution of mixotrophy in the Orchideae orchid tribe, and of ectomycorrhizal abilities in the Ceratobasidiaceae. Our new ectomycorrhiza formation technique may enhance the study of unculturable orchid mycorrhizal fungi.     

Phylogeny, diversity, and classification of the Accipitridae based on DNA sequences of the RAG-1 exon

The avian family Accipitridae has historically been divided into subfamilies or tribes based on features such as general resemblance, feeding ecology, and behavior. Consequently, the monophyly of those groups has been questionable. Recently, three phylogenetic analyses of a majority of the genera have appeared, one based on osteology, one on DNA sequences from a single mitochondrial gene, and the third on mitochondrial plus nuclear DNA sequences, and the resulting phylogenies were in substantial disagreement concerning the composition and basal branching patterns of the clades and hence require further analysis and confirmation. Here we use DNA sequences from the large nuclear RAG-1 exon to investigate the phylogenetic relationships of these birds. Our results largely corroborated the prior study that included nuclear genes. We found strong support for a monophyletic clade comprising the secretarybird Sagittarius serpentarius , the osprey Pandion haliaetus , and the traditional accipitrids. However, every one of the traditionally recognized subfamilies of accipitrids was found to be polyphyletic. The most basal nodes in the phylogeny separate small clades of insectivorous and scavenger species, such as kites and Old World vultures, from the rest of the family. The speciose genera of bird and mammal predators are all relatively derived (terminal) in the phylogeny. Many of the basal clades are cosmopolitan in their distributions, consistent with the great mobility of these raptors. A new classification is proposed that eliminates the problem of polyphyletic intrafamilial taxa.     

Molecular analysis of orchid pollinaria and pollinaria-remains found on insects

Direct observations of pollinator visits to orchids are often difficult and time consuming, especially in orchids with a deceptive pollination system where seed set is typically pollinator-limited. This lack of direct observations greatly inhibits our understanding of orchid-pollinator relationships and especially the degree of pollinator-specificity. Here we describe a molecular approach to the study of orchid-pollinator relationships based on the analysis of DNA recovered from pollinaria found on insects. The insects were collected from nectar-rich plants flowering near natural orchid populations, or taken from museum collections. Sequence analysis of the nuclear ribosomal ITS region allowed the identification of the orchid species or species-group from which the pollinaria originated. Four out of eight orchid-pollinator relationships established with this approach have not been reported previously, which highlights the value of molecular tools for the study of orchid pollination biology.     

A complete generic phylogeny of Malpighiaceae inferred from nucleotide sequence data and morphology

? Premise of the study: The Malpighiaceae include ～1300 tropical flowering plant species in which generic definitions and intergeneric relationships have long been problematic. The goals of our study were to resolve relationships among the 11 generic segregates from the New World genus Mascagnia, test the monophyly of the largest remaining Malpighiaceae genera, and clarify the placement of Old World Malpighiaceae. ? Methods: We combined DNA sequence data for four genes (plastid ndhF, matK, and rbcL and nuclear PHYC) from 338 ingroup accessions that represented all 77 currently recognized genera with morphological data from 144 ingroup species to produce a complete generic phylogeny of the family. ? Key results and conclusions: The genera are distributed among 14 mostly well-supported clades. The interrelationships of these major subclades have strong support, except for the clade comprising the wing-fruited genera (i.e., the malpighioid+Amorimia, Ectopopterys, hiraeoid, stigmaphylloid, and tetrapteroid clades). These results resolve numerous systematic problems, while others have emerged and constitute opportunities for future study. Malpighiaceae migrated from the New to Old World nine times, with two of those migrants being very recent arrivals from the New World. The seven other Old World clades dispersed much earlier, likely during the Tertiary. Comparison of floral morphology in Old World Malpighiaceae with their closest New World relatives suggests that morphological stasis in the New World likely results from selection by neotropical oil-bee pollinators and that the morphological diversity found in Old World flowers has evolved following their release from selection by those bees.     

Large multi-gene phylogenetic trees of the grasses (Poaceae): progress towards complete tribal and generic level sampling

In this paper we included a very broad representation of grass family diversity (84% of tribes and 42% of genera). Phylogenetic inference was based on three plastid DNA regions rbcL, matK and trnL-F, using maximum parsimony and Bayesian methods. Our results resolved most of the subfamily relationships within the major clades (BEP and PACCMAD), which had previously been unclear, such as, among others the: (i) BEP and PACCMAD sister relationship, (ii) composition of clades and the sister-relationship of Ehrhartoideae and Bambusoideae + Pooideae, (iii) paraphyly of tribe Bambuseae, (iv) position of Gynerium as sister to Panicoideae, (v) phylogenetic position of Micrairoideae. With the presence of a relatively large amount of missing data, we were able to increase taxon sampling substantially in our analyses from 107 to 295 taxa. However, bootstrap support and to a lesser extent Bayesian inference posterior probabilities were generally lower in analyses involving missing data than those not including them. We produced a fully resolved phylogenetic summary tree for the grass family at subfamily level and indicated the most likely relationships of all included tribes in our analysis.     

Strong phylogenetic congruence between Tulasnella fungi and their associated Drakaeinae orchids

The study of congruency between phylogenies of interacting species can provide a powerful approach for understanding the evolutionary history of symbiotic associations. Orchid mycorrhizal fungi can survive independently of orchids making cospeciation unlikely, leading us to predict that any congruence would arise from host-switches to closely related fungal species. The Australasian orchid subtribe Drakaeinae is an iconic group of sexually deceptive orchids that consists of approximately 66 species. In this study, we investigated the evolutionary relationships between representatives of all six Drakaeinae orchid genera (39 species) and their mycorrhizal fungi. We used an exome capture dataset to generate the first well-resolved phylogeny of the Drakaeinae genera. A total of 10 closely related Tulasnella Operational Taxonomic Units (OTUs) and previously described species were associated with the Drakaeinae orchids. Three of them were shared among orchid genera, with each genus associating with 1–6 Tulasnella lineages. Cophylogenetic analyses show Drakaeinae orchids and their Tulasnella associates exhibit significant congruence (p < 0.001) in the topology of their phylogenetic trees. An event-based method also revealed significant congruence in Drakaeinae–Tulasnella relationships, with duplications (35), losses (25), and failure to diverge (9) the most frequent events, with minimal evidence for cospeciation (1) and host-switches (2). The high number of duplications suggests that the orchids speciate independently from the fungi, and the fungal species association of the ancestral orchid species is typically maintained in the daughter species. For the Drakaeinae–Tulasnella interaction, a pattern of phylogenetic niche conservatism rather than coevolution likely explains the observed phylogenetic congruency in orchid and fungal phylogenies. Given that many orchid genera are characterized by sharing of fungal species between closely related orchid species, we predict that these findings may apply to a wide range of orchid lineages.