Sara B. Hoot

Inferring Complex Phylogenies Using Parsimony: An Empirical Approach Using Three Large DNA Data Sets for Angiosperms

To explore the feasibility of parsimony analysis for large data sets, we conducted heuristic parsimony searches and bootstrap analyses on separate and combined DNA data sets for 190 angiosperms and three outgroups. Separate data sets of 18S rDNA (1,855 bp), rbcL (1,428 bp), and atpB (1,450 bp) sequences were combined into a single matrix 4,733 bp in length. Analyses of the combined data set show great improvements in computer run times compared to those of the separate data sets and of the data sets combined in pairs. Six searches of the 18S rDNA + rbcL + atpB data set were conducted; in all cases TBR branch swapping was completed, generally within a few days. In contrast, TBR branch swapping was not completed for any of the three separate data sets, or for the pairwise combined data sets. These results illustrate that it is possible to conduct a thorough search of tree space with large data sets, given sufficient signal. In this case, and probably most others, sufficient signal for a large number of taxa can only be obtained by combining data sets. The combined data sets also have higher internal support for clades than the separate data sets, and more clades receive bootstrap support of > or = 50% in the combined analysis than in analyses of the separate data sets. These data suggest that one solution to the computational and analytical dilemmas posed by large data sets is the addition of nucleotides, as well as taxa.

Phylogenetic Relationships of Capsicum (Solanaceae) Using DNA Sequences from Two Noncoding Regions: The Chloroplast atpB‐rbcL Spacer Region and Nuclear waxy Introns

This study focuses on three phylogenetic problems related to Capsicum (Solanaceae): (1) the monophyly of the genus, (2) species delimitation within the genus, and (3) phylogenetic relationships of species within Capsicum. The chloroplast atpB‐rbcL noncoding spacer region was used to derive a phylogeny for seven outgroup genera and 11 species of Capsicum. Data derived from five introns within the nuclear gene waxy were used, both separately and in combination with the atpB‐rbcL spacer data, to resolve further questions of species delimitation and phylogenetic relationships within Capsicum. Capsicum is monophyletic, with moderate support. Capsicum ciliatum, which is both molecularly and morphologically distinctive, is sister to a highly supported clade consisting of all other Capsicum species studied. Capsicum cardenasii and C. eximium are sister species and are, in turn, sisters to a moderately supported clade consisting of C. tovarii, C. pubescens, C. chacoense, C. baccatum, C. galapagoense, C. chinense, C. frutescens, and C. annuum. Capsicum galapagoense, whose taxonomic affinities have been largely unstudied, is included in a weakly supported clade consisting of C. annuum, C. chinensis, and C. frutescens. Many species of Capsicum have sufficient molecular markers in the waxy data set (both nucleotide substitutions and insertions/deletions) to be useful in species delimitation. An informal classification of the genus is proposed.

Patterns of floral evolution in the early diversification of non-magnoliid dicotyledons (eudicots)

Recent cladistic analyses of angiosperms based on both morphological and molecular sequence data recognize a major clade of dicotyledons defined by triaperturate or triaperturate-derived pollen (non-magnoliids/eudicots). Evidence from morphology, as well as the atpB and rbcL genes (cpDNA), indicates that extant Ranunculidae (e.g., Papaverales, Lardizabalaceae, Berberidaceae, Menispermaceae, Ranunculaceae) as well as “lower” Hamamelididae [e.g., Eupteleaceae (allied to Ranunculidae), Hamamelidaceae, Myrothamnaceae, Platanaceae, Trochodendraceae] and several other families (e.g., Gunneraceae, Nelumbonaceae, Proteaceae, Sabiaceae) are basal in this group. The earliest records of diagnostic eudicot pollen are of mid-late Barremian age (c. 126myr BP) and by around the latest Albian (c. 97 myr BP) several basal eudicot groups (e.g., Trochodendrales, Platanaceae, Buxaceae, and perhaps Circaeasteraceae, Myrothamnaceae, and Nelumbonaceae) are recognizable in the fossil record. Possible Hamamelidaceae and perhaps Proteaceae are present by the Turonian (c. 90 myr BP). Among basal eudicots, flowers are generally bisexual although unisexual flowers are also common. In some groups (e.g., Myrothamnaceae, Buxaceae, certain Berberidaceae), delimitation of the flower is not always clear and there is a more or less gradual transition between tepals and inflorescence bracts. Plasticity in floral form at this level of angiosperm evolution is predominantly encompassed by dimerous and trimerous cyclic floral organization and transitions from one to the other are common. Spiral floral phyllotaxis of numerous stamens and carpels is more or less restricted to the Ranunculaceae. The basic condition of the perianth in eudicots appears to lack differentiation into sepals and petals, and petals appear to have arisen independently numerous times from stamens. Based on the generality of its systematic distribution, cyclic floral architecture is probably basic for eudicots as a whole, and at this level of angiosperm evolution flowers with numerous, helically-arranged stamens and/or carpels (e.g., many Ranunculaceae) almost certainly reflect processes of secondary multiplication that have occurred independently many times.

Phylogeny of the Ranunculaceae based on preliminary atpB, rbcL and 18S nuclear ribosomal DNA sequence data

The chloroplast genes atpB and rbcL and nuclear ribosomal 18S DNA were sequenced for 23 genera of the Ranunculaceae and two outgroup taxa (Hydrastis and Glaucidium). The three sequence data sets were combined and the resulting preliminary phylogenetic tree used to assess relationships within the Ranunculaceae. The phylogeny strongly supports the monophyly of the family, with 26 substitutions, a bootstrap value of 98% and a decay index of > 7. Within the family, the T-type chromosome group is basal and paraphyletic with respect to the larger R-type chromosome group. Within the T-type chromosome group, Coptis and Xanthorhiza from a monophyletic group and are basal to all other Ranunculaceae. Other alliances previously proposed by taxonomists are confirmed: Anemonella/Thalictrum/ lsopyrum/Aquilegia/Semiaquilegia; Anemone/Clematis, Trautvetteria/ Myosurus/Ranunculus; Aconitum/Delphinium; and Anemonopsis/Cimicifugal/Actaea. Other groupings that could not have been predicted on the basis of traditional data include a clade consisting of Adonis and Trollius and the inclusion of Eranthis in a clade with Anemonopsis, Cimicifuga, and Actaea. Nigella is weakly allied with a clade consisting of Aconitum and Delphinium. The molecular sequence data are largely congruent with results based on cytology, phytochemistry, and micromorphology. Flower and fruit characters are homoplastic in relation to the cladogram based on sequence data. The pattern of relationships based on sequence data supports the view that staminodia/petals and achenes have evolved independently several times within the Ranunculaceae.

PHYLOGENETICS OF THE HAMAMELIDAE AND THEIR ALLIES: PARSIMONY ANALYSES OF NUCLEOTIDE SEQUENCES OF THE PLASTID GENE rbcL

Over the past decade it has become increasingly clear that the angiosperm subclass Hamamelidae is polyphyletic. To examine phylogenetic affinities of various lineages in the complex and to evaluate the positions of lower hamamelids in angiosperm phylogeny, we performed a parsimony analysis using nucleotide sequences of the plastid gene rbcL from 134 species, including representatives of 25 families that have traditionally been placed in the Hamamelidae; most rosid families; and selected members of the caryophyllids, asterids, monocots, and magnoliids. Platanaceae, Eupteleaceae, Tetracentraceae, Trochodendraceae, Buxaceae, Didymeleaceae, and Myrothamnaceae occupy basal positions in the eudicots, together with Ranunculales‐Papaverales, Nelumbonaceae, Proteaceae, and Gunneraceae. Cercidiphyllaceae, Daphniphyllaceae, and Hamamelidaceae are placed among basal rosids, being closely related to Paeonia and Saxifragaceae s.s. and their allies. Nothofagaceae, Fagaceae, Juglandaceae, Ticodendraceae, Myricaceae, Betulaceae, and Casuarinacae form a monophyletic group, which in some trees is embedded in a large rosid clade that includes many nitrogen‐fixing species. Ulmaceae, Moraceae, Cannabaceae, Urticaceae, Cecropiaceae, and Barbeyaceae are also part of this “nitrogen‐fixing clade,” and they are related to Rhamnus, Elaeagnus, Dirachma, and Rosaceae. Leitneriaceae are clearly a member of Sapindales. Eucommiaceae form a clade with Aucuba and Garrya among the asterids.

The Utility of Nuclear ITS, a LEAFY Homolog Intron, and Chloroplast atpB-rbcL Spacer Region Data in Phylogenetic Analyses and Species Delimitation in Isoetes

Abstract Despite its ancient origins, its worldwide distribution, and adaptation to diverse habitats, Isoëtes has a highly conserved morphology. This feature has made it difficult to resolve species and species relationships using morphological characters. In this paper, we report the utility of nucleotide sequences from the nuclear internal transcribed spacer (ITS) regions, chloroplast atpB/rbcL intergenic spacer region, and second intron of a LEAFY (LFY) homolog for identifying species relationships, delimiting basic diploid species, and determining hybrid origins. Variation in the ITS regions and atpB/rbcL spacer is most useful at the family level in Isoëtes and the LFY second intron is appropriate at the species and population level. The tree resulting from an analysis of the combined nuclear ITS and chloroplast atpB/rbcL spacer contains three major well supported clades (bootstrap ≥ 99%): an Old-World/California clade (I. abyssinica, I. longissima, I. velata, I. nuttallii, and I. orcuttii), an Asian/Australian clade (I. taiwanensis, I. japonica, I. kirkii, and I. drummondii), and a poorly resolved clade consisting of nine North American species. To further resolve and delimit the North American species, a combination of the LEAFY second intron and ITS data was used. The resulting consensus tree has limited resolution, supporting the hypothesis that the North American species complex radiated rapidly. The combination of LFY and ITS data provided numerous characters, both substitutions and indels, that are useful in species delimitation and identification of cryptic species. ITS sequence data, through additive banding and sequence misalignment, is also useful in confirming interspecific hybrids and determining their parental origins.

Data congruence and phylogeny of the Papaveraceae s.l. based on four data sets: atpB and rbcL sequences, trnK restriction sites, and morphological characters

Phylogenetic analyses of various combinations of atpB and rbcL sequence data, trnK restriction sites, and morphological characters of taxa representing the Papaveraceae s.l. (Pteridophyllaceae, Fumari- aceae subfamilies Fumarioideae and Hypecoideae, Papaveraceae s. str. subfamilies Eschscholzioideae, Chelidonioideae, Papaveroideae, Platystemonoideae) are presented. Several measures of data set incongru- ency indicate considerable congruence between data sets. In order to correctly identify the root of the family, combined atpB and rbcL sequence data for the Papaveraceae s.l. and appropriate outgroups (Asarum, Illicium, Platanus, Euptelea, and representatives of the Ranunculaceae and Berberidaceae) were analyzed. Regardless of the outgroup used, all topologies obtained suggest that Pteridophyllum (Pteridophyllaceae) is sister group to Fumariaceae and Papaveraceae s. str. Data sets of the ingroup with Pteridophyllum used as the outgroup were analyzed separately and together, with results from the combined data sets showing increased bootstrap support. Contrary to recent claims, Papaveraceae s. str. remain monophyletic after the exclusion of Pteridophyl- lum and Fumariaceae. In the Papaveraceae s. str., Eschscholzioideae are sister group to Papaveroideae (including Platystemonoideae) and Chelidonioideae (including Glaucium/Dicranostigma). The evolution of floral morphology, geographical distribution, and ecology of Papaveraceae s.l. are briefly discussed.

Phylogeny of the Ranunculaceae based on epidermal microcharacters and macromorphology

The epidermal microcharacters of 20 genera in the Ranunculaceae and 10 outgroup genera from the families Glaucidiaceae, Circaeasteraceae, Berberidaceae, Menispermaceae, and Lar- dizabalaceae were studied employing scanning electron and light microscopy. Characters such as type of trichome and stomatal length are valuable tools in assessing phylogeny in the Ranunculaceae. Using cladistic analyses, the basic division of the family into two major groups according to chro- mosome size (R- and T-types) was confirmed. By outgroup polarization, the T-type chromosome group is considered most primitive. Two major clades are recognized in the T-type taxa: the Aquilegia and Thalictrum group including closely related genera and a group consisting of Coptis and Xan- thorhiza. Hydrastis is not consjdered a member of the Ranunculaceae. Further cladistic analyses of the R-type taxa using macrocharacters derived mainly from floral and fruit morphology resulted in the following subdivisions: the Anemone and Ranunculus group including Pulsatilla, Hepatica, and Clematis; the Cimicifuga and Actaea group; and the Helleborus group including Eranthis, Trollius, and Caltha. The Ranunculaceae, a large and varied fam- ily, consist of approximately 66 genera and 2000 species (Takhtajan 1987). Members of the family are distributed throughout the world, but are centered in the temperate and cold regions of both hemispheres; representatives are uncom- mon in tropical and subtropical zones except in montane regions (Ziman and Keener 1989).

Inter-familial relationships in the Ranunculidae based on molecular systematics

Nucleotide sequences from the rbcL and atpB genes (chloroplast DNA) and 18S nuclear ribosomal DNA provide important new data with which to test previous hypotheses of inter-familial relationships in the angiosperm subclass Ranunculidae. Preliminary conclusions based on cladistic analysis of the combined molecular data sets for all three genes are broadly congruent with previous analyses based on rbcL data alone. The genus Euptelea, which has been placed traditionally among the “lower” Hamamelididae, is resolved as more closely related to ranunculids. Papaverales (represented by Pteridophyllaceae and Fumariaceae including Hypecoum) are strongly supported as a monophyletic group, and are resolved as basal to all other ranunculids (“core” ranunculids) plus Euptelea. The monotypic genera Kingdonia and Circaeaster are placed as sister taxa and together are sister to Lardizabalaceae s.l. (including Sargentodoxa). The Lardizabalaceae-Circaeaster-Kingdonia clade is the sister taxon to Menispermaceae-Ranunculaceae-Berberidaceae. Menispermaceae is the sister taxon to Ranunculaceae and Berberidaceae.

Systematics and phylogeny of the Ranunculaceae — a revised family concept on the basis of molecular data

Phylogenetic trees for the Ranunculaceae based on four independent molecular data sets are compared and analyzed. This comparison includes chloroplast DNA restriction site variation; atpB (chloroplast), rbcL, nuclear ribosomal DNA sequences (analysed as a combined data set), nuclear adh sequences, and serological detected characters of the major seed protein, legumin. These trees are highly congruent in terminal branching patterns with high support. This congruency suggests a strong correlation between the evolution of the genes and proteins investigated and the taxa involved and further demonstrates the utility of molecular markers in plant phylogeny. The molecular results are often congruent with non-molecular data as well and are used to develop more reliable systematic classification of the Ranunculaceae.