Mark E. Mort

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.

Phylogenetics and Evolution of the Macaronesian Clade of Crassulaceae Inferred from Nuclear and Chloroplast Sequence Data

Abstract The Macaronesian clade of Crassulaceae comprises four genera (Aichryson, Aeonium, Greenovia, and Monanthes) that are largely endemic to Macaronesia, a region encompassing the Azores, Madeira, and the Cape Verde, Canary, and Salvage Islands. The monophyly of this clade has been supported by recent family-level phylogenetic analyses; however, the relationships within the clade remain uncertain. To resolve relationships within the Macaronesian clade, we sequenced several chloroplast DNA regions (matK and the trnL-trnF and psbA-trnH spacer regions) and the nuclear rDNA ITS region. Parsimony analyses of separate ITS and cpDNA data sets recover three major clades, corresponding to Aichryson, the perennial species of Monanthes, and Aeonium (including Greenovia). Congruence tests revealed significant heterogeneity between the ITS and cpDNA data sets. Comparison of the topologies resulting from analyses of these separate data sets indicated five instances of incongruence between the ITS and cpDNA that may be the result of cpDNA capture events. Those five cpDNA sequences that appear to be the result of hybridization were removed, and a combined cpDNA/ITS data set was constructed and analyzed. Parsimony analyses of this combined data set again resolve three major clades that correspond to Aichryson, Monanthes, and Aeonium (including Greenovia); however, the combined analyses provide greater resolution and higher internal support than the analyses of the separate data sets. Given this estimate of phylogeny, the Macaronesian clade most likely evolved from herbaceous, continental ancestors; the woody habit evolved once in Aichryson tortuosum and again in the ancestor of the Aeonium clade. Combined analyses of cpDNA and ITS for a large sampling of the Macaronesian Crassulaceae also confirms recent conclusions that Aeonium originated in Macaronesia and not in Africa and that the east African species, A. leucoblepharum, is of recent origin and represents long-distance dispersal from Macaronesia to Africa. Communicating Editor: Paul Wilson

Inferring phylogeny at low taxonomic levels: utility of rapidly evolving cpDNA and nuclear ITS loci

Plant molecular systematic studies of closely related taxa have relied heavily on sequence data from nuclear ITS and cpDNA. Positive attributes of using ITS sequence data include the rapid rate of evolution compared to most plastid loci and availability of universal primers for amplification and sequencing. On the other hand, ITS sequence data may not adequately track organismal phylogeny if concerted evolution and high rDNA array copy number do not permit identification of orthologous copies. Shaw et al. (American Journal of Botany 92: 142-166) recently identified nine plastid regions that appear to provide more potentially informative characters than many other plastid loci. In the present study, sequences of these loci and ITS were obtained for six taxonomic groups in which phylogenetic relationships have been difficult to establish using other data. The relative utility of these regions was compared by assessing the number of parsimony informative characters, character congruence, resolution of inferred trees, clade support, and accuracy. No single locus emerged as the best in all lineages for any of these measures of utility. Results further indicated that in preliminary studies, sampling strategy should include at least four exemplar taxa. The importance of sampling data from independent distributions is also discussed.

Timing and Tempo of Early and Successive Adaptive Radiations in Macaronesia

The flora of Macaronesia, which encompasses five Atlantic archipelagos (Azores, Canaries, Madeira, Cape Verde, and Salvage), is exceptionally rich and diverse. Spectacular radiation of numerous endemic plant groups has made the Macaronesian islands an outstanding area for studies of evolution and speciation. Despite intensive investigation in the last 15 years, absolute age and rate of diversification are poorly known for the flora of Macaronesia. Here we report molecular divergence estimates and rates of diversification for five representative, putative rapid radiations of monophyletic endemic plant lineages across the core eudicot clade of flowering plants. Three discrete windows of colonization during the Miocene and early Pliocene are suggested for these lineages, all of which are inferred to have had a single colonization event followed by rapid radiation. Subsequent inter-archipelago dispersal events into Madeira and the Cape Verdes took place very recently during the late Pliocene and Pleistocene after initial diversification on the Canary Islands. The tempo of adaptive radiations differs among the groups, but is relatively rapid compared to continental and other island radiations. Our results demonstrate that opportunity for island colonization and successful radiation may have been constrained to discrete time periods of profound climatic and geological changes in northern African and the Mediterranean.

Elucidating deep-level phylogenetic relationships in Saxifragaceae using sequences for six chloroplastic and nuclear DNA regions

To elucidate relationships at deep levels within Saxifragaceae we analyzed phylogenetically a data set of sequences for six DNA regions, four representing the chloroplast genome (rbcL, matK, trnL-trnF, psbA-trnH) and two from the nuclear genome (ITS and expansion segments of the 26S rDNA). A total of 6676 bp was aligned per taxon, 4559 bp and 1878 bp from the chloroplast and nuclear genomes, respectively. Chloroplast and nuclear trees agreed closely, prompting analysis of a combined, six-gene data set. Application of both parsimony and maximum likelihood methods yielded similar topologies. The use of different ITS alignments and the exclusion of hard-to-align ITS regions had little impact on either the final nuclear-based topology, or the shortest trees from the analysis of six genes. The affinities of two monotypic genera (Saxifragella and Saxifragodes) endemic to Tierra del Fuego were elucidated. Saxifragella is an early branching member of the North Temperate genus Saxifraga s. str.; Saxifragodes is sister to Cascadia, a genus endemic to Oregon and Washington. Long-distance dispersal from east Asia or western North America to South America may have played an important role in forming these and other similar disjunctions in the family. A number of well-supported clades are present, including Saxifraga s. str., Micranthes, Saxifragopsis/Astilbe, Chrysosplenium/Peltoboykinia, and the Boykinia and Heuchera groups. The use of additional characters has provided greatly increased resolution and internal support at deep levels. Saxifragaceae comprise two major lineages: Saxifraga s. str. (including Saxifragella) and all other genera of the family (the heucheroids). This major split is accompanied by general biogeographical and morpihological differences. Whereas Saxifraga s. str. is largely arctic to alpine in occurrence, the heucheroid clade is largely temperate in distribution, Saxifraga s. str. has a relatively uniform floral morphology (generally actinomorphic; 5 sepals, 5 petals, 10 stamens, 2 carpels), whereas the heucheroid clade encompasses actinomorphic and zygomorphic forms, as well as variation in the number of sepals, petals, stamens, and carpels. Deep-level relationships within both Saxifraga s. str. and the heucheroid clade are well resolved and supported. A phylogenetic classification of the family is provided.

Direct estimation of the mutation rate at dinucleotide microsatellite loci in Arabidopsis thaliana (Brassicaceae).

The mutation rate at 54 perfect (uninterrupted) dinucleotide microsatellite loci is estimated by direct genotyping of 96 Arabidopsis thaliana mutation accumulation lines. The estimated rate differs significantly among motif types with the highest rate for AT repeats (2.03 × 10−3 per allele per generation), intermediate for CT (3.31 × 10−4), and lowest for CA (4.96 × 10−5). The average mutation rate per generation for this sample of loci is 8.87 × 10−4 (s.e.=2.57 × 10−4). There is a strong effect of initial repeat number, particularly for AT repeats, with mutation rate increasing with the length of the microsatellite locus in the progenitor line. Controlling for motif and initial repeat number, chromosome 4 exhibited an elevated mutation rate relative to other chromosomes. The great majority of mutations were gains or losses of a single repeat. Generally, the data are consistent with the stepwise mutation model of microsatellite evolution. Several lines exhibited multiple step changes from the progenitor sequence, but it is unclear whether these are multi-step mutations or multiple single-step mutations. A survey of dinucleotide repeats across the entire Arabidopsis genome indicates that AT repeats are most abundant, followed by CT, and CA.

The utility of automated analysis of inter-simple sequence repeat (ISSR) loci for resolving relationships in the Canary Island species of Tolpis (Asteraceae)

Plants of oceanic islands, often remarkably divergent morphologically from continental relatives, are useful models for studying evolution and speciation because evolution is telescoped in time and space. Prior studies revealed little DNA sequence variation within the clade of ca. 10 Canary Island species of Tolpis, which precluded resolving species relationships. The present study assessed the utility of automated analysis of inter-simple sequence repeat (ISSR) loci for resolving relationships within the clade using 264 individuals from 36 populations of all recognized species and three undescribed morphological variants. Similarity (Dice coefficient) and Fitch parsimony were used to generate neighbor-joining (NJ) and strict consensus trees (MP), respectively. All individuals of the morphologically distinct endemic species formed clusters in both trees. There is also support for clusters of two undescribed variants in the NJ tree. Individuals from a morphologically variable complex consisting primarily of two species are not well resolved at population or species levels. The NJ and MP trees are not congruent at deeper levels, including relationships among species. Results are interpreted in terms of the biology of the species, and the utility of automated analysis of ISSR markers for interpreting patterns of evolution of Tolpis in the Canary Islands is discussed.

Phylogenetic relationships and character evolution analysis of Saxifragales using a supermatrix approach

UNLABELLED PREMISE OF THE STUDY We sought novel evolutionary insights for the highly diverse Saxifragales by constructing a large phylogenetic tree encompassing 36.8% of the species-level biodiversity. • METHODS We built a phylogenetic tree for 909 species of Saxifragales and used this hypothesis to examine character evolution for annual or perennial habit, woody or herbaceous habit, ovary position, petal number, carpel number, and stamen to petal ratio. We employed likelihood approaches to investigate the effect of habit and life history on speciation and extinction within this clade. • KEY RESULTS Two major shifts occurred from a woody ancestor to the herbaceous habit, with multiple secondary changes from herbaceous to woody. Transitions among superior, subinferior, and inferior ovaries appear equiprobable. A major increase in petal number is correlated with a large increase in carpel number; these increases have co-occurred multiple times in Crassulaceae. Perennial or woody lineages have higher rates of speciation than annual or herbaceous ones, but higher probabilities of extinction offset these differences. Hence, net diversification rates are highest for annual, herbaceous lineages and lowest for woody perennials. The shift from annuality to perenniality in herbaceous taxa is frequent. Conversely, woody perennial lineages to woody annual transitions are infrequent; if they occur, the woody annual state is left immediately. • CONCLUSIONS The large tree provides new insights into character evolution that are not obvious with smaller trees. Our results indicate that in some cases the evolution of angiosperms might be conditioned by constraints that have been so far overlooked.

Relationships among the Macaronesian members of Tolpis (Asteraceae: Lactuceae) based upon analyses of inter simple sequence repeat (ISSR) markers

With the advent of PCR and DNA sequencing, DNA data have been widely applied to the assessment of relationships among taxa of recent origin (e.g., island endemics). Numerous recent studies have used DNA sequence data and/or cpDNA RFLPs to reconstruct phylogeny for many members of the Macaronesian flora. One such genus that has received attention recently is Tolpis (Asteraceae: Lactuceae). Sequences of the cpDNA gene ndhF provided strong support for a clade comprising species of Tolpis that are largely endemic to Macaronesia, to the exclusion of several European taxa that had previously been placed within the genus. Subsequently, cpDNA RFLP data were used to assess the biogeography and potential patterns of dispersal within the genus. However, relationships among species of Tolpis present on the Canary Islands were not well-resolved. Analyses of intersimple sequence repeat (ISSR) data were employed to assess relationships among species of Tolpis from the Canary Islands and Madeira. The five primers used across 80 individuals from 10 populations of seven species resulted in 48 bands that could be scored consistently. Neighbor-joining and parsimony analyses indicate close relationships among individuals within populations and differentiation among allopatric conspecific populations. In agreement with prior molecular results, the Canarian species form a well-supported group. Both neighbor-joining and parsimony analyses of ISSR markers resolve relationships (often with bootstrap support) among species, something that was not achieved previously. The present study demonstrates the potential of hypervariable single-locus markers for elucidating relationships in rapidly radiating lineages, many of which occur in oceanic archipelagos.

A TEST OF BAKER'S LAW: BREEDING SYSTEMS AND THE RADIATION OF TOLPIS (ASTERACEAE) IN THE CANARY ISLANDS

Baker’s law posits that self‐compatible (SC) plants will be more successful than self‐incompatible (SI) plants in long‐distance colonization because a single propagule can establish a viable population. Oceanic islands represent ideal systems to test Baker’s law because insular lineages have, without question, originated from long‐distance dispersal. The dilemma of Baker’s law is that one propagule of an SC plant would establish a population with low genetic diversity, which could limit subsequent evolution. By contrast, a single propagule from an SI ancestor, having originated from an outcrossing source population, would provide more diversity but could not undergo sexual reproduction. We examined this issue by studying the breeding system of members of the flowering plant genus Tolpis (Asteraceae), a small (nine to 13 species), monophyletic lineage in the Canary Islands archipelago. A combination of floral morphology, pollen‐ovule ratio, autogamous seed set, and genetic data indicates that only one endemic species (T. coronopifolia) is effectively SC. The remainder of the endemics are pseudo‐self‐compatible, i.e., are largely SI but capable of low levels of seed set from self‐fertilization. Pseudo‐self‐compatibility remedies the dilemma of Baker’s law: a single propagule can establish a sexual population and yet have sufficient variation to facilitate diversification.