Phylogeny of the freshwater crabs of the Western Ghats (Brachyura,Gecarcinucidae)

The Western Ghats mountain range in India is a biodiversity hotspot for a variety of organisms including a large number of endemic freshwater crab species and genera of the family Gecarcinucidae. The phylogenetic relationships of these taxa, however, have remained poorly understood. Here, we present a phylogeny that includes 90% of peninsular Indian genera based on mitochondrial 16S rRNA and nuclear histone H3 gene sequences. The subfamily Gecarcinucinae was found to be paraphyletic with members of two other subfamilies, Liotelphusinae and Parathelphusinae, nesting within. We identify a well‐supported clade consisting of north Indian species and one clade comprising mostly south Indian species that inhabit the southern ‘sky islands’ of the Western Ghats. Relationships of early diverging genera, however, were resolved with low support. This study also includes newly sampled material from an isolated mountain plateau in the northern part of the Western Ghats, representing a new species of Gubernatoriana, which we describe here as Gubernatoriana basalticola sp. n. The new species is immediately distinguished from its congeners and the related genera Ghatiana and Inglethelphusa by its carapace and cheliped morphology, which are unique among Indian freshwater crabs. This study highlights the urgent need for continued faunistic studies to assess the true diversity of gecarcinucid crabs on the Indian subcontinent, to fully understand the basal phylogenetic relationships within the freshwater crab family Gecarcinucidae, and to evaluate the conservation threat status and biogeography of the montane freshwater crabs of the Western Ghats.     

Fly pollination in Ceropegia (Apocynaceae: Asclepiadoideae): biogeographic and phylogenetic perspectives

Background and Aims

Ceropegia (Apocynaceae subfamily Asclepiadoideae) is a large, Old World genus of >180 species, all of which possess distinctive flask-shaped flowers that temporarily trap pollinators. The taxonomic diversity of pollinators, biogeographic and phylogenetic patterns of pollinator exploitation, and the level of specificity of interactions were assessed in order to begin to understand the role of pollinators in promoting diversification within the genus.

Methods

Flower visitor and pollinator data for approx. 60 Ceropegia taxa were analysed with reference to the main centres of diversity of the genus and to a cpDNA–nrDNA molecular phylogeny of the genus.

Key Results

Ceropegia spp. interact with flower-visiting Diptera from at least 26 genera in 20 families, of which 11 genera and 11 families are pollinators. Size range of flies was 0·5–4·0 mm and approx. 94 % were females. Ceropegia from particular regions do not use specific fly genera or families, though Arabian Peninsula species are pollinated by a wider range of Diptera families than those in other regions. The basal-most clade interacts with the highest diversity of Diptera families and genera, largely due to one hyper-generalist taxon, C. aristolochioides subsp. deflersiana. Species in the more-derived clades interact with a smaller diversity of Diptera. Approximately 60 % of taxa are so far recorded as interacting with only a single genus of pollinators, the remaining 40 % being less conservative in their interactions. Ceropegia spp. can therefore be ecological specialists or generalists.

Conclusions

The genus Ceropegia has largely radiated without evolutionary shifts in pollinator functional specialization, maintaining its interactions with small Diptera. Intriguing biogeographic and phylogenetic patterns may reflect processes of regional dispersal, diversification and subsequent specialization onto a narrower range of pollinators, though some of the findings may be caused by inconsistent sampling. Comparisons are made with other plant genera in the Aristolochiaceae and Araceae that have evolved flask-shaped flowers that trap female flies seeking oviposition sites.Key words: Apocynaceae, Asclepiadoideae, Brachystelma, Ceropegia, Diptera, flower evolution, generalization, mutualism, pollination, Riocreuxia, specialization, Stapeliinae     

Polyphyletic origin in <Emphasis Type="Italic">Pimpinella</Emphasis> (Apiaceae): evidence in Western Europe

The genus Pimpinella L. comprises about 150 species, being one of the largest genera within the family Apiaceae (subfamily Apioideae). Previous molecular phylogenetic studies have shown that Pimpinella is a taxonomically complex group. In this study, evolutionary relationships among representatives from Western Europe have been inferred from phylogenetic analyses of nuclear ribosomal DNA internal transcribed spacer (ITS 1 and ITS 2) and plastid sequences (trnL intron and the trnL-F spacer), with a representative sampling included (168 accessions in the ITS analysis, representing 158 species; and 42 accessions in the cpDNA analysis representing 35 taxa of Pimpinella and closely related species). All analyses resolved that Pimpinella is a non-monophyletic group, and Pimpinella’s taxa that grow in Western Europe are part of phylogenetically independent groups that correspond to three different tribes of the subfamily Apioideae: Pimpinelleae (core group), Pyramidoptereae and Smyrnieae.     

Origin and diversification of Indian Ceropegieae (Apocynaceae) and its possible relation to the Indian monsoon

The Indian subcontinent has experienced a major shift in climatic regime from a wet tropical regime to increased seasonal rainfall, since the late Miocene. This shift has been attributed to the intensification of monsoons, which led to opening up of dry habitats in humid forests and formation of deciduous forests. We explored the role of this climatic shift in the origin and diversification of dry‐adapted plant genera Ceropegia and Brachystelma (Ceropegiae, Asclepiadoideae, Apocynaceae). We sampled Ceropegia and Brachystelma from across India and used five markers (two nuclear and three plastid regions) to reconstruct a global phylogeny of this group. Indian members of the tribe Ceropegiae were derived from Africa through at least four independent dispersal events. All dispersal events occurred in late Miocene after establishment of a monsoon climate. One of these early dispersing lineages underwent rapid radiation in peninsular India, giving rise to around 50 species. Thus, both dispersal and diversification events coincided with the intensification of monsoons and concomitant aridification. The role of environment in the evolution of floral characteristics and root type in the Indian radiation is also discussed. This is one of the first reports on a dry‐adapted endemic radiation of plants in India.     

a molecular phylogeny of apiaceae subfamily Apioideae: evidence from nuclear ribosomal DNA internal transcribed spacer sequences

Phylogenetic relationships among 40 New World and Old World members of Apiaceae subfamily Apioideae, representing seven of the eight tribes and eight of the ten subtribes commonly recognized in the subfamily, were inferred from nucleotide sequence variation in the internal transcribed spacer (ITS) regions of 18-26S nuclear ribosomal DNA. Although the sequences are alignable, with only 11% of sites excluded from the analyses because of alignment ambiguity, divergence values in pairwise comparisons of unambiguous positions among all taxa were high and ranged from 0.5 to 33.2% of nucleotides in ITS 1 and from 0 to 33.2% of nucleotides in ITS 2. Average sequence divergence across both spacer regions was 18.4% of nucleotides. Phylogenies derived from ITS sequences estimated using neighbor-joining analysis of substitution rates, and maximum likelihood and parsimony methods give trees of essentially similar topology and indicate that: (1) there is little support for any existing system of classification of the subfamily that is based largely on morphological and anatomical features of the mericarp; (2) there is a major phylogenetic division within the subfamily, with one clade comprising the genus Smyrnium and those taxa belonging to Drude's tribes Dauceae, Scandiceae, and Laserpitieae and the other clade comprising all other examined taxa; and (3) the genera Arracacia, Coaxana, Coulterophytum, Enantiophylla, Myrrhidendron, Prionosciadium, and Rhodosciadium, all endemic to Mexico and Central America, comprise a clade but their relationships to other New World taxa are equivocal. A phylogeny derived from parsimony analysis of chloroplast DNA rpoC1 intron sequences is consistent with, but considerably less resolved than, relationships derived from these ITS regions. This study affirms that ITS sequences are useful for phylogenetic inference among closely related members of Apioideae but, owing to high rates of nucleotide substitution, are less useful in resolving relationships among the more ancestral nodes of the phylogeny.     

DISCORDANCE BETWEEN NUCLEAR AND CHLOROPLAST PHYLOGENIES IN THE HEUCHERA GROUP (SAXIFRAGACEAE)

Various factors, including taxon density, sampling error, convergence, and heterogeneity of evolutionary rates, can potentially lead to incongruence between phylogenetic trees based on different genomes. Particularly at the generic level and below, chloroplast capture resulting from hybridization may distort organismal relationships in phylogenetic analyses based on the chloroplast genome, or genes included therein. However, the extent of such discord between chloroplast DNA (cpDNA) trees and those trees based on nuclear genes has rarely been assessed. We therefore used sequences of the internal transcribed spacer regions (ITS-1 and ITS-2) of nuclear ribosomal DNA (rDNA) to reconstruct phylogenetic relationships among members of the Heuchera group of genera (Saxifragaceae). The Heuchera group presents an important model for the analysis of chloroplast capture and its impact on phylogenetic reconstruction because hybridization is well documented within genera (e.g., Heuchera), and intergeneric hybrids involving six of the nine genera have been reported. An earlier study provided a well-resolved phylogenetic hypothesis for the Heuchera group based on cpDNA restriction-site variation. However, trees based on ITS sequences are discordant with the cpDNA-based tree. Evidence from both morphology and nuclear-encoded allozymes is consistent with the ITS trees, rather than the cpDNA tree, and several points of phylogenetic discord can clearly be attributed to chloroplast capture. Comparison of the organellar and ITS trees also raises the strong likelihood that ancient events of chloroplast capture occurred between lineages during the early diversification of the Heuchera group. Thus, despite the many advantages and widespread use of cpDNA data in phylogeny reconstruction, comparison of relationships based on cpDNA and ITS sequences for the Heuchera group underscores the need for caution in the use of organellar variation for retrieving phylogeny at lower taxonomic levels, particularly in groups noted for hybridization.     

Phylogenetic reticulation in subtribe Helianthinae

Incongruence between phylogenetic estimates based on nuclear and chloroplast DNA (cpDNA) markers was used to infer that there have been at least two instances of chloroplast transfer, presumably through wide hybridization, in subtribe Helianthinae. One instance involved Simsia dombeyana, which exhibited a cpDNA restriction site phenotype that was markedly divergent from all of the other species of the genus that were surveyed but that matched the restriction site pattern previously reported for South American species of Viguiera. In contrast, analysis of sequence data from the nuclear ribosomal DNA internal transcribed spacer (ITS) region showed Simsia to be entirely monophyletic and placed samples of S. dombeyana as the sister group to the relatively derived S. foetida, a result concordant with morphological information. A sample of a South American species of Viguiera was placed by ITS sequence data as the sister group to a member of V. subg. Amphilepis, which was consistent with cpDNA restriction site data. Samples of Tithonia formed a single monophyletic clade based on ITS sequence data, whereas they were split between two divergent clades based on cpDNA restriction site analysis. The results suggested that cpDNA transfer has occurred between taxa diverged to the level of morphologically distinct genera, and highlight the need for careful and complete assessment of molecular data as a source of phylogenetic information.     

Variation in the nrDNA ITS of Pinus Subsection Cembroides: Implications for Molecular Systematic Studies of Pine Species Complexes

The pinyon pines (Pinus subsection Cembroides), distributed in semiarid regions of the western United States and Mexico, include a mixture of relictual and more recently evolved taxa. To investigate relationships among the pinyons, we screened and partially sequenced 3000-bp clones of the nuclear ribosomal DNA internal transcribed spacer (ITS) region for 16 taxa from subsect. Cembroides and nine representatives from four other subsections of subgenus Strobus. Restriction digests of clones reveal within-individual heterogeneity, suggesting that concerted evolution is operating slowly on the ITS in pine species. Two ITS clones were identified as pseudogenes. Tandem subrepeats in the ITS1 form stem loops comparable to those in other genera of Pinaceae and may be promoting recombination between rDNA repeats, resulting in ITS1 chimeras. Within the pinyon clade, phylogenetic structure is present, but different clones from the same (or different) individuals of a species are polyphyletic, indicating that coalescence of ITS copies within individual genomes predates evolutionary divergence in the group. At the level of subsection and above, the ITS region corresponds well with morphological and cpDNA evidence. Except for P. nelsonii, the pinyons are monophyletic, with both subsect. Cembroides and P. nelsonii forming a clade with the foxtail and bristlecone pines (subsect. Balfourianae) of western North America.     

The molecular phylogeny of aquatic hyphomycetes with affinity to the Leotiomycetes

Aquatic hyphomycetes play a key role in decomposition of submerged organic matter and stream ecosystem functioning. We examined the phylogenetic relationships among various genera of aquatic hyphomycetes belonging to the Leotiomycetes (Ascomycota) using sequences of internal transcribed spacer (ITS) and large subunit (LSU) regions of rDNA generated from 42 pure cultures including 19 ex-types. These new sequence data were analyzed together with additional sequences from 36 aquatic hyphomycetes and 60 related fungi obtained from GenBank. Aquatic hyphomycetes, characterized by their tetraradiate or sigmoid conidia, were scattered in nine supported clades within the Helotiales (Leotiomycetes). Tricladium, Lemonniera, Articulospora, Anguillospora, Varicosporium, Filosporella, and Flagellospora are not monophyletic, with species from the same genus distributed among several major clades. The Gyoerffyella clade and the Hymenoscyphus clade accommodated species from eight and six different genera, respectively. Thirteen aquatic hyphomycete taxa were grouped in the Leotia-Bulgaria clade while twelve species clustered within the Hymenoscyphus clade along with several amphibious ascomycetes. Species of Filosporella and some species from four other aquatic genera were placed in the Ascocoryne-Hydrocina clade. It is evident that many aquatic hyphomycetes have relatives of terrestrial origin. Adaptation to colonize the aquatic environment has evolved independently in multiple phylogenetic lineages within the Leotiomycetes.     

Phylogenetic relationship in Fritillaria spp. of Iran inferred from ribosomal ITS and chloroplast trnL-trnF sequence data

The genus Fritillaria embraces up to 165 taxa in the family Liliaceae, most of which are of high medicinal and ornamental value and importance. In this study, 44 specimens of the genus representing 9 species were collected from their natural habitats located in 10 provinces of Iran. Phylogenetic analysis was performed based on DNA sequences of the internal transcribed spacer (ITS) of the nuclear ribosomal cistron and the trnL-trnF regions. The phylogeny was constructed using the neighbor joining inference method. Results indicate that the examined samples were evidently diverged into 2 distinct clades. Members of the subgenera Fritillaria and Rhinopetalum formed one clade while the other clade contained the subgenera Theresia and Petilium. There can be seen a high degree of similarity between the only yellow-colored crown imperial specimen and the red-colored specimens. The endemic species of Fritillaria straussii, Fritillaria zagrica and Fritillaria kotschyana which their status within the subgenera known in the genus Fritillaria has been remained undefined, fell into the subgenus Fritillaria. The clades also had relatively reasonable distribution patterns based on the genetic structure, geographical conditions and climate specifications. This study revealed the feasibility of the ITS and trnL-trnF DNA sequence for phylogeny of the genus Fritillaria. This is the first phylogenetic analysis of Fritillaria spp. in Iran.     

A phylogenetic analysis ofPanax (Araliaceae): Integrating cpDNA restriction site and nuclear rDNA ITS sequence data

A phylogenetic analysis ofPanax was performed using restriction site variations of eight PCR-amplified chloroplast regions. Twenty populations were examined, representing 13 of the 14 species ofPanax. Aralia cordata was used as the outgroup. The 11 restriction endonucleases produced a total of 105 restriction sites and length variations from the large single-copy region of cpDNA. Forty restriction variations are polymorphic. The cpDNA tree is largely congruent with the nuclear ribosomal ITS phylogeny. Similar to the ITS tree, the cpDNA dataset suggests the following relationships: (1)P. trifolius from eastern North America is sister to the clade consisting of all otherPanax species; (2)P. ginseng andP. japonicus from eastern Asia form a clade withP. quinquefolius from eastern North America; (3) the HimalayanP. pseudoginseng is most closely related toP. stipuleanatus of southwestern China; and (4) the medicinally importantP. notoginseng forms a clade with the closely relatedP. bipinnatifidus, P. ginseng, P. japonicus, P. major, P. quinquefolius, P. sinensis, P. wangianus, andP. zingiberensis. Two biogeographic disjunctions are detectable withinPanax. One is the connection of the eastern North AmericanP. trifolius with the rest ofPanax species. The other is the more recent disjunction between the North AmericanP. quinquefolius and the eastern AsianP. ginseng andP. japonicus. The active orogenies caused by the collision of the Indian Plate with Asia may have facilitated the diversification ofPanax taxa in Asia in the late Tertiary.     

Nuclear ribosomal ITS sequences and phylogeny in East AsianAconitum subgenusAconitum (Ranunculaceae), with special reference to extensive polymorphism in individual plants

Internal transcribed spacer (ITS) sequences of nuclear ribosomal DNA (nrDNA) were used to examine the phylogeny of East Asian aconites. Individual aconites were discovered to contain as many as eight different ITS sequences after cloning and PCR-SSCP (single-stranded conformational polymorphisms) analysis. We identified eight putative ITS pseudogenes from four taxa with low predicted secondary structure stability and high substitution rates. Maximum likelihood (ML) and neighbor-joining (NJ) methods were used for phylogenetic reconstruction. The ITS trees agree with the previous chloroplast DNA (cpDNA) tree for the vast majority of the taxa. We found two East Asian clades in the ITS trees: 1) a clade with the Chinese diploid,Aconitum volubile and East Asian tetraploids, and 2) a clade of East Asian diploids and Siberian tetraploids. In the former clade, most tetraploid taxa appear to be polyphyletic; sequences from individual plants did not correspond to recognized taxonomic units. This indicates a recent divergence of the East Asian tetraploids.     

Phylogenetic split of Larix: evidence from paternally inherited cpDNA trnT-trnF region

A molecular phylogeny of Larix comprising 12 species was constructed from the sequence analysis of the paternally inherited cpDNA trnT-trnF region of 46 individuals. The most parsimonious tree split Larix into three sister clades: one clade was composed of two North American species, the other two were short-bracted and long-bracted species of Eurasia respectively except that L. sibirica was clustered in the long-bracted clade. The difference between the present cpDNA phylogeny and previous nrDNA ITS phylogeny in the position of L. sibirica seems to suggest that ancient cytoplasmic gene flow might exist between sections Larix and Multiserialis. The short-bracted L. laricina and long-bracted L. occidentalis have an identical sequence of the trnT-trnF region, which implies that the bract length divergence among North American larches might have occurred recently or chloroplast capture happened during the early differentiation of the two species. The cpDNA results also shed some light on the biogeography of Larix.     

Evidence of chloroplast capture in South American Nothofagus (subgenus Nothofagus,Nothofagaceae)

Subgenus Nothofagus, although geographically restricted at present to temperate areas of South America, has captured much attention in discussions of plant biogeography due to its widespread distribution through Gondwanan continents during the Tertiary. However, phylogenetic relationships within the subgenus Nothofagus have not yet been resolved. We examined geographic patterns of intraspecific and interspecific genetic variation to detect whether incongruences in nuclear or plastid DNA phylogenies occur, in order to better understand the evolutionary history of the subgenus Nothofagus. We conducted spatially-explicit sampling at 10 distinct locations throughout the range of austral South American forests and sampled all present Nothofagus species. We used ITS and chloroplast DNA sequences to estimate phylogenetic relationships. A phylogeny constructed from nuclear genes resolved the subgenus Nothofagus as monophyletic. We found that N. antarctica was a sister to a clade of evergreen species (N. betuloides, N. dombeyi, and N. nitida), while N. pumilio likely diverged earlier. Nine cpDNA haplotypes were distinguished in the subgenus Nothofagus which were associated to geographic locations rather than to taxonomic relationships. This species-independent cpDNA phylogeographic structures within the subgenus Nothofagus may be related to repeated chloroplast capture events over geological time in Patagonia.     

Variation in the nrDNA ITS of Pinus subsection Cembroides: implications for molecular systematic studies of pine species complexes.

The pinyon pines (Pinus subsection Cembroides), distributed in semiarid regions of the western United States and Mexico, include a mixture of relictual and more recently evolved taxa. To investigate relationships among the pinyons, we screened and partially sequenced 3000-bp clones of the nuclear ribosomal DNA internal transcribed spacer (ITS) region for 16 taxa from subsect. Cembroides and nine representatives from four other subsections of subgenus Strobus. Restriction digests of clones reveal within-individual heterogeneity, suggesting that concerted evolution is operating slowly on the ITS in pine species. Two ITS clones were identified as pseudogenes. Tandem subrepeats in the ITS1 form stem loops comparable to those in other genera of Pinaceae and may be promoting recombination between rDNA repeats, resulting in ITS1 chimeras. Within the pinyon clade, phylogenetic structure is present, but different clones from the same (or different) individuals of a species are polyphyletic, indicating that coalescence of ITS copies within individual genomes predates evolutionary divergence in the group. At the level of subsection and above, the ITS region corresponds well with morphological and cpDNA evidence. Except for P. nelsonii, the pinyons are monophyletic, with both subsect. Cembroides and P. nelsonii forming a clade with the foxtail and bristlecone pines (subsect. Balfourianae) of western North America.     

A molecular phylogeny and generic rearrangement of the stapelioid Ceropegieae (Apocynaceae-Asclepiadoideae)

 Representatives of nearly all genera of the taxon-rich stem-succulent stapeliads and most of the few related, leafy genera were analyzed. Sequence data from two non-coding molecular markers (ITS region of nrDNA and trnT-L and trnL-F spacers as well as the trnL intron of cpDNA) support the traditional tribal affiliation of the genera, which form a monophyletic group. This monophylum breaks into a basal Neoschumannia/Anisotoma/Riocreuxia/Sisyranthus nk;clade, from which the core Ceropegieae are derived. The four Ceropegia species included are not monophyletic, and their relationship to Brachystelma changes depending on the marker studied. The stem succulent taxa fall in a number of well supported, but unresolved clades, the most prominent being the predominantly southern African clade comprising Orbea, Stapelia and some other genera. The most derived taxa of NE Africa, Duvaliandra and White-sloanea, are basal to this southern African clade. The other clades comprise the more basal genera of stem-succulent stapeliads, including the members of the Caralluma complex. Of the 17 genera accepted by Plowes for the Caralluma complex, seven are recognized: Caralluma, Apteranthes, Australluma, Boucerosia, Caudanthera, Desmidorchis and Monolluma. New combinations are proposed in 15 cases; Caralluma adscendens var. geniculata is raised to specific rank. Anomalluma is reinstated, and Pseudolithos mccoyi is transfered to it. A broadened concept for Orbea (incl. Angolluma and Orbeopsis) is recognized, but Orbeanthus is kept separate. The monotypic Ballyanthus, recently separated from Orbea, is nested within Duvalia. Piaranthus (incl. Huerniopsis) is monophyletic. The bitypic Notechidnopsis is reduced to the type species, N. tessellata, while N. columnaris is transferred to a new genus, Richtersveldia. Received February 25, 2002; accepted June 17, 2002 Published online: November 7, 2002 Address of the authors: Dr. Ulrich Meve (e-mail: ulrich.meve@uni-bayreuth.de) and Prof. Dr. Sigrid Liede (e-mail: sigrid.liede@uni-bayreuth.de), Universit?t Bayreuth, Lehrstuhl für Pflanzensystematik, Universit?tsstrasse 30, D-95440 Bayreuth, Germany.     

A molecular phylogeny shows the single origin of the Pyrenean subterranean Trechini ground beetles (Coleoptera: Carabidae)

Trechini ground beetles include some of the most spectacular radiations of cave and endogean Coleoptera, but the origin of the subterranean taxa and their typical morphological adaptations (loss of eyes and wings, depigmentation, elongation of body and appendages) have never been studied in a formal phylogenetic framework. We provide here a molecular phylogeny of the Pyrenean subterranean Trechini based on a combination of mitochondrial (cox1, cyb, rrnL, tRNA-Leu, nad1) and nuclear (SSU, LSU) markers of 102 specimens of 90 species. We found all Pyrenean highly modified subterranean taxa to be monophyletic, to the exclusion of all epigean and all subterranean species from other geographical areas (Cantabrian and Iberian mountains, Alps). Within the Pyrenean subterranean clade the three genera (Geotrechus, Aphaenops and Hydraphaenops) were polyphyletic, indicating multiple origins of their special adaptations to different ways of life (endogean, troglobitic or living in deep fissures). Diversification followed a geographical pattern, with two main clades in the western and central-eastern Pyrenees respectively, and several smaller lineages of more restricted range. Based on a Bayesian relaxed-clock approach, and using as an approximation a standard mitochondrial mutation rate of 2.3% MY, we estimate the origin of the subterranean clade at ca. 10 MY. Cladogenetic events in the Pliocene and Pleistocene were almost exclusively within the same geographical area and involving species of the same morphological type.