An ITS-based phylogenetic framework for the genus Vorticella: finding the molecular and morphological gaps in a taxonomically difficult group

Vorticella includes more than 100 currently recognized species and represents one of the most taxonomically challenging genera of ciliates. Molecular phylogenetic analysis of Vorticella has been performed so far with only sequences coding for small subunit ribosomal RNA (SSU rRNA); only a few of its species have been investigated using other genetic markers owing to a lack of similar sequences for comparison. Consequently, phylogenetic relationships within the genus remain unclear, and molecular discrimination between morphospecies is often difficult because most regions of the SSU rRNA gene are too highly conserved to be helpful. In this paper, we move molecular systematics for this group of ciliates to the infrageneric level by sequencing additional molecular markers—fast-evolving internal transcribed spacer (ITS) regions—in a broad sample of 66 individual samples of 28 morphospecies of Vorticella collected from Asia, North America and Europe. Our phylogenies all featured two strongly supported, highly divergent, paraphyletic clades (I, II) comprising the morphologically defined genus Vorticella. Three major lineages made up clade I, with a relatively well-resolved branching order in each one. The marked divergence of clade II from clade I confirms that the former should be recognized as a separate taxonomic unit as indicated by SSU rRNA phylogenies. We made the first attempt to elucidate relationships between species in clade II using both morphological and multi-gene approaches, and our data supported a close relationship between some morphospecies of Vorticella and Opisthonecta, indicating that relationships between species in the clade are far more complex than would be expected from their morphology. Different patterns of helix III of ITS2 secondary structure were clearly specific to clades and subclades of Vorticella and, therefore, may prove useful for resolving phylogenetic relationships in other groups of ciliates.     

PHYLOGENY OF THE EUGLENOID LORICATE GENERA TRACHELOMONAS AND STROMBOMONAS (EUGLENOPHYTA) INFERRED FROM NUCLEAR SSU AND LSU rDNA1

Previous studies using the nuclear SSU rDNA and partial LSU rDNA have demonstrated that the euglenoid loricate taxa form a monophyletic clade within the photosynthetic euglenoid lineage. It was unclear, however, whether the loricate genera Trachelomonas and Strombomonas were monophyletic. In order to determine the relationships among the loricate taxa, SSU and LSU nuclear rDNA sequences were obtained for eight Strombomonas and 25 Trachelomonas strains and combined in a multigene phylogenetic analysis. Conserved regions of the aligned data set were used to generate maximum‐likelihood (ML) and Bayesian phylogenies. Both methods recovered a strongly supported monophyletic loricate clade with Strombomonas and Trachelomonas species separated into two sister clades. Taxa in the genus Strombomonas sorted into three subclades. Within the genus Trachelomonas, five strongly supported subclades were recovered in all analyses. Key morphological features could be attributed to each of the subclades, with the major separation being that all of the spine‐bearing taxa were located in two sister subclades, while the more rounded, spineless taxa formed the remaining three subclades. The separation of genera and subclades was supported by 42 distinct molecular signatures (33 in Trachelomonas and nine in Strombomonas). The morphological and molecular data supported the retention of Trachelomonas and Strombomonas as separate loricate genera.     

Phylogenetic relationships within the aquatic snail genus Tryonia: implications for biogeography of the North American Southwest

We examined the phylogenetic relationships among 23 species of the North American aquatic snail genus Tryonia (Hydrobiidae), 10 additional representatives of the subfamily Cochliopinae, and two outgroups. Maximum parsimony analysis of a 601-base-pair sequence from the mitochondrial COI gene did not support monophyly of the genus nor its subgenus Paupertryonia. A clade composed of the type species of Tryonia and 16 congeners was strongly supported by the COI data and congruent with recently discovered variation in female genitalic morphology. This "true Tryonia" clade included two large western subclades having a sister-group relationship. The phylogenetic structure of one of these subclades is congruent with vicariant events associated with late Neogene history of the lower Colorado River drainage. The other subclade mirrors development of the modern Rio Grande rift and inception of modern topography in the southwestern Great Basin during the late Neogene. Both subclades are represented in the composite Tryonia fauna of the Amargosa River basin, whose assembly is attributed to the complex geological history of the Death Valley region.     

Molecular phylogenetic analysis of Euphorbiaceae sensu stricto based on plastid and nuclear DNA sequences and ovule and seed character evolution

A phylogenetic analysis of Euphorbiaceae sensu stricto is presented using sequences from rbcL, atpB, matK and 18S rDNA from 85 species and 83 genera. The combined analysis of four molecular markers resulted in only one most parsimonious tree and also generated new supported clades, which include Euphorbioideae + Acalyphoideae s.s., subclades A2 + A3, subclades A5 + A6 and a clade uniting subclades A2–A8 within Acalyphoideae s.s. A palisadal exotegmen is a possible synapomorphy for all the Euphorbiaceae, except for the subfamily Peroideae. The presence of vascular bundles in the inner integument and a thick inner integument were shown to be synapomorphies for the clade of inaperturate and articulated crotonoids and for the large clade of Euphorbioideae, Acalyphoideae s.s., inaperturate and articulated crotonoids, respectively. Characters of the aril and vascular bundles in the outer integument are discussed. The selected embryological characters were seen to be highly correlated with the molecular phylogeny. When the results of molecular phylogenetic analysis of a previous study and this study were adjusted along with the selected embryological characters, all clades within Euphorbiaceae were supported except for a clade comprising Euphorbioideae + Acalyphoideae s.s. + inaperturate crotonoids + articulated crotonoids + Adenoclineae s.l. and a clade uniting subclades A4–A8 within Acalyphoideae s.s. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

中国南蛇藤属 (卫矛科) 一新记录种及其意义

在开展南蛇藤属分类修订的过程中,于 2011 年在中国云南省西双版纳傣族自治州勐腊县发现南蛇藤属中国分布新记录——拟独子藤 ( Celastrus monospermoides) 。该种形态与独子藤 ( C. monospermus) 较为相似,主要区别为前者叶片窄椭圆形,先端渐尖,果实成熟后不具雌蕊柄,种子长12 mm 以内; 后者叶片椭圆形、阔椭圆形至倒卵椭圆形,果实具有明显的雌蕊柄,种子长15 mm 以上。分子系统学研究表明,拟独子藤与独子藤处于不同分支中,与青江藤 ( C. hindsii) 系统关系最近。由于原记录仅分布于印度尼西亚、马来西亚和菲律宾等热带地区,因此在原产地和我国云南省之间的地区应有更多居群分布,这为探讨南蛇藤属的起源和扩散提供了线索。     

Delimitation and phylogeny of Aletris (Nartheciaceae) with implication for perianth evolution

Aletris, containing approximately 21 species, is the largest genus in Nartheciaceae, and is disjunctively distributed in eastern Asia and eastern North America. Its delimitation has been controversial because it is uncertain whether Metanarthecium should be included in the genus. Although there are a few molecular phylogenetic studies on Aletris, the interspecific relationships within the genus have never been evaluated in a phylogenetic context. Here we used two cpDNA loci, matK and trnL-F, to delimitate Aletris and discuss the phylogeny within the genus. Phylogenetic analyses showed Metanarthecium might be distantly related to Aletris. This is also supported by morphological, palynological, cytological, and phytochemical data. Therefore, Metanarthecium should be excluded from Aletris. Within Aletris, there are two major clades: A. farinosa and A. lutea of eastern North America and A. glabra of eastern Asia form clade A; and the remaining Asian species form clade B. The Asian clade includes three subclades: subclade I (two varieties of A. pauciflora, and A. glandulifera and A. megalantha), subclade II (three samples of A. laxiflora), and subclade III (all other sampled Asian species). Based on phylogenetic relationships, A. pauciflora var. khasiana deserves a specific status, and A. gracilipes, formerly a synonym of A. laxiflora, should be reinstated. The reconstruction of the perianth evolution indicates that perianth connate halfway and glabrous on abaxial surface are plesiomorphic for Aletris and Nartheciaceae. Farinose-glutinous perianth is a diagnostic character for clade A.     

Patterns of coral-dinoflagellate associations in Acropora: significance of local availability and physiology of Symbiodinium strains and host-symbiont selectivity

Like other reef-building corals, members of the genus Acropora form obligate endosymbioses with dinoflagellates (zooxanthellae) belonging to the genus Symbiodinium. Both Symbiodinium and its hosts are diverse assemblages, and the relationships between host and algal genotypes are unclear. In this study, we determined phylogenetic relationships between Symbiodinium isolates from a wide range of Acropora species and plotted the algal genotypes onto a molecular phylogeny of 28 Acropora species, using the same samples for the host and symbiont genotyping. In addition, we performed a preliminary survey of zooxanthella distribution in Acropora species from the central Great Barrier Reef. Three of the four known major zooxanthellae clades were represented in the 168 samples examined, and within the major clade C, three distinct subclades were identified. No evidence was found for coevolution, but several clear patterns of specificity were identified. Moreover, composition of the zooxanthella pool varied among locales and in one host species we found light-related patterns of zooxanthella distribution.     

Phylogeny of the Neotropical Alibertia group (Rubiaceae), with emphasis on the genus Alibertia, inferred from ITS and 5S ribosomal DNA sequences

Phylogenetic relationships of 38 species of the Alibertia group (Rubiaceae) and two outgroup species were investigated using the nuclear ribosomal 5S nontranscribed spacer (5S-NTS) and the internal transcribed spacers (ITS). Analysis of the data sets separately and in combination resulted in several well-supported and congruent groupings. However, the three analyses yielded different results as to the branching order of the basal clades. With the exception of Alibertia hispida, the species in the genus Alibertia appear in one weakly to moderately supported clade. This clade is in turn composed of two strongly supported subclades. One comprises several Alibertia species, including the type (A. edulis), three Borojoa species, and Randia tessmannii. The other subclade consists of Alibertia species only. This division is also generally supported morphologically by fruit size, corolla size, number of corolla lobes, and pollen aperture (porate vs. colporate). The sister group to the Alibertia clade comprises Duroia with Amaioua species internested. The close relationship of Ibetralia and Kutchubaea is corroborated. In addition, Alibertia hispida is a member of this strongly supported clade. Likewise, the two "Genipa" species are supported as a monophyletic group in 100% of the bootstrap replicates. It is concluded that the 5S spacer is superior to the commonly used ITS region in terms of resolution and robustness among closely related taxa.     

Phylogenetic position of South American Cheilanthes (Cheilanthoideae,Pteridaceae): Advances in the generic circumscription and segregation of the new genus Mineirella

This analysis corroborates and expands our previous results regarding the phylogenetic position of Cheilanthes species from South America. We sequenced three plastid genetic regions, one genic (rbcL) and two genic plus intergenic spacers (trnL + trnL-F and rps4 + rps4-trnS) from 25 South American cheilanthoid species. This allowed us to elucidate phylogenetic relationships that have been historically unresolved or were lowly supported. Here, we analyzed 45 Cheilanthes species (23 from South America) and circumscribed Cheilanthes s.s. in a strongly supported clade that contains three subclades: (i) exclusively from South America, (ii) from Australasia + South America, and (iii) from Africa. The position of three South American species, previously referred to the informal “Cheilanthes geraniifolia group”, is confirmed as a highly supported group outside Cheilanthes s.s. and within the Adiantopsis–Doryopteris clade. This group is described here as the new genus Mineirella. The new combinations for the genus and illustrations are included. Additionally, we discuss the morphological innovations that provide evidence to support the different clades.     

碎米蕨属的系统学研究——基于4种叶绿体DNA序列片段

碎米蕨属(Cheilosoria Trevis.)隶属碎米蕨类, 由于形态上的趋同进化, 使得该类群的系统分类一直存在争议。该研究对该碎米蕨属植物的4种叶绿体DNA序列片段(rbcL/matK/rps4/rps4-trnS)进行PCR扩增和序列分析, 再结合其它相关类群, 用贝叶斯法和最大似然法构建系统树并探讨其系统发育关系。结果表明, 碎米蕨属不是一个单系类群, 旧世界分布的碎米蕨属植物(薄叶碎米蕨除外)均聚在亚洲Cheilanthes群内, 与粉背蕨属(Aleuritopteris Fée)等类群形成不同亚支。该属孢子形态具有明显异质性。薄叶碎米蕨(Cheilosoria tenuifolia (Burm. f.) Trevis.)与亚洲其它碎米蕨属植物的系统位置相距甚远, 且与隐囊蕨(Notholaena hirsuta (Poir.) Desv.)聚为完全支持的分支, 两者可能均为大洋洲起源, 并属于另一类群。美洲和旧世界分布的碎米蕨属植物关系较远, 二者可能代表了不同的演化路线。     

Molecular studies of photobionts of selected lichens from the coastal vegetation of Brazil

A light microscopic and molecular analysis of photobionts in Ramalina and Cladonia from coastal habitats of Brazil is presented. A Bayesian phylogenetic analysis of ITS rDNA sequences suggests a Trebouxia lineage which is preferentially tropical in geographic distribution. This highly diverse clade also includes the morphological similar species Trebouxia higginsiae and galapagensis. Within the predominantly tropical clade of Trebouxia we distinguish several subclades, three of which are represented in our samples of Ramalina species. Since sexuality has not been recognized in coccal lichenised photobionts until recently, we cannot apply a biological species concept, but when compared with the sequence diversity between known species we conclude that several new species need to be described in this clade. The mutually exclusive presence of other Trebouxia lineages in temperate samples of Ramalina suggests an evolution towards higher selectivity in this genus. A strictly tropical lineage is not conspicuous in the photobionts of the genus Asterochloris sampled from Cladonia so far.     

Phylogenetic relationships of the anamorphic genus <Emphasis Type="Italic">Fusicladium</Emphasis> s. lat. as inferred by ITS nrDNA data

The genus Fusicladium s. lat. (incl. Pollaccia and Spilocaea) was phylogenetically analysed using ITS nrDNA sequences. Pollaccia and Spilocaea did not form monophyletic groups of their own, but were intermingled between Fusicladium species, together with which they formed a monophyletic clade. Thus, Pollaccia and Spilocaea should be included in a wider genus concept of Fusicladium, constituting a morphologically variable genus. Furthermore, all Venturia and Fusicladium isolates clustered together on the bases of available ITS data, providing support for the monophyly of the anamorphic genus Fusicladium and the teleomorphic genus Venturia. Within this clade several subclades can be recognized. All taxa on the host family Salicaceae were found in one subclade. Three other subclades comprised taxa on Rosaceae whereas taxa on other host families all clustered separately. Geographic specializations were not observed. Two examples of host switching could be demonstrated, but these were confined to instances involving host species belonging to the same family. Fusicladium convolvularum and F. effusum, two species with unknown teleomorphs, clustered within the Fusicladium/Venturia clade, supporting the correct placement of these taxa in Fusicladium. The placement of Pseudocladosporium hachijoense within the family Venturiaceae was also supported.     

中国桫椤科植物叶绿体trnL内含子和trnL-trnF基因间隔区序列的系统发育分析

运用对PCR产物直接测序和克隆后测序的方法测定了蚌壳蕨科1种和桫椤科11种(其中桫椤分别测定19株：小羽桫椤测定2株)植物的叶绿体trnL基因内含子和trnL-trnF基因间隔区序列。12种植物相应的长度介亍l004-l082之间,A T平均含量60．9％,G C平均含量39．1％。计算了不同种间以及种内不同个体间序列的碱基差别(转换值／颠换值)和Kimura遗传距离。序列数据经排列后分别进行最简约法、最大似然法和邻接法分析,结果显示：(1)白桫椤、海南白桫椤和大羽桫椤构成的分支最早和该科内其余植物组成的另一分支分歧,而后者又进一步分为刃个亚分支,分别和桫椤亚属、黑桫椤亚属对应,支持夏群的分类处理：(2)大桫椤～狭羽桫椤～毛轴桫椤～篦齿桫椤、多羽桫椤一白桫椤～海南白桫椤以及小羽桫椤一桫椤各自构成独立、自然的末端分支,再参照分支内植物间的遗传距蔼取值,建议将此3个末端分支依次归并为3种：大桫椤、白桫椤和桫椤;(3)白桫椤属在科内处于基部位置,桫椤属奇桫椤亚属和黑桫椤亚属为衍生分支,赞同Tryon关于桫椤科进化和囊群盖起源的假说。     

Molecular phylogeny of terrestrial holocarpic endoparasitic peronosporomycetes, <Emphasis Type="Italic">Haptoglossa</Emphasis> spp., inferred from 18S rDNA

Phylogenetic relationships of seven isolates of the genus Haptoglossa parasitic on terrestrial nematodes within the Peronosporomycetes were analyzed using 18S rDNA sequence data with 21 peronosporomycetes, 2 marine stramenopilous flagellates, and 2 hyphochytridiomycetes. The marine stramenopilous flagellates and hyphochytridiomycetes were used as the outgroup. All Haptoglossa isolates formed a monophyletic clade and clustered with the marine genus Eurychasma. The clade of Haptoglossa and Eurychasma formed a sister-group to the clade that consisted of all other peronosporomycetes. These results suggest that the genus Haptoglossa and other terrestrial peronosporomycetes included in the two subclasses, the Saprolegniomycetidae and the Peronosporomycetidae, might have originally adapted to the terrestrial environment individually. In the maximum-likelihood (ML) analysis, the Haptoglossa clade was divided into three subclades, one aplanosporic species clade and two zoosporic species clades. Phylogenetic analyses of combined 18S rDNA and cox2 genes among five species of Haptoglossa supported the results of the ML analysis using 18S rDNA and suggested that zoosporic species may be separated into two lineages. This topology of the analysis may suggest that aplanosporic species diverged from zoosporic species.     

Molecular support for a rapid cladogenesis of the woodpecker clade Malarpicini, with further insights into the genus Picus (Piciformes: Picinae)

Previous studies have suggested that the woodpecker genus Picus (Aves: Picidae) may not be monophyletic. In order to evaluate this hypothesis, we analyzed DNA sequences from all but two species of Picus, as well as from representatives of all genera in the tribe Malarpicini, within which Picus is nested. We sequenced seven loci (four autosomal, one Z-linked and two mitochondrial) with different evolutionary dynamics. The species currently placed in Picus fall into two subclades that may not form a monophyletic assemblage. Consequently, we propose to place miniaceus Pennant 1769, flavinucha Gould 1834 and mentalis Temminck 1825 in the genus Chrysophlegma Gould, 1850, while the remaining species are retained in Picus. The inclusion in our study of representatives of all genera included in the tribe Malarpicini, a group of woodpeckers which has proven difficult to resolve in several previous molecular studies, also allowed us to determine the earliest divergences within this clade. The results suggest that the low level of basal resolution in Malarpicini is attributable to multiple cladogenetic events in a short period of time rather than insufficient character sampling. This conclusion is supported by the observation of nucleotide insertion-deletions that support mutually exclusive phylogenetic hypotheses in different gene trees. We attribute this pattern of incongruent indels, together with short internodes in the tree, to incomplete lineage sorting.     

Molecular evolutionary relationships in the avian genus Anthus (Pipits: Motacillidae)

Nucleotide sequences for 1035 bp of the mitochondrial cytochrome b gene were used to determine the molecular evolutionary relationships of species in the cosmopolitan avian genus Anthus. Phylogenetic analysis of these mtDNA sequences supported four major clades within the genus: (1) the small-bodied African pipits, (2) a largely Palearctic clade, (3) a largely South American clade, and (4) an African-Eurasian-Australian clade. Anthus hellmayri, A. correndera, and A. rubescens are shown to be paraphyletic. The possibility of paraphyly within A. similis is instead inferred to be the discovery of a new species and supported by reference to the museum voucher specimen. Sequence divergence suggests a Pliocene/Miocene origin for the genus. Although Anthus cytochrome b is found not to be behaving in a clocklike fashion across all taxa, speciation during the Pleistocene epoch can be reasonably inferred for the 66% of sister pairs that are diverging in a clocklike manner. Base compositions at each codon position are similar to those found across a growing number of avian lineages. The resulting phylogenetic hypothesis is compared to previous hypotheses of Anthus relationships, all of which deal with relationships of a particular species or a particular species complex; roughly half of these previous hypotheses are supported.     

A nuclear DNA phylogeny and proposed taxonomic revision of African greenbuls (Aves, Passeriformes, Pycnonotidae)

The Pycnonotidae (bulbuls and greenbuls) comprise approximately 130 species and are widely distributed across Africa and Asia, mainly in evergreen thickets and forest. Recent molecular findings suggest a basal split between the African and the Asian species, although the three African Pycnonotus species are part of the Asian radiation and represent a relative recent immigration to Africa. In this study we investigate the phylogenetic relationships within the African clade, which with the exclusion of Pycnonotus contains approximately 50 species, of which the majority are placed in three large genera Andropadus , Phyllastrephus and Chlorocichla . We use three nuclear markers (myoglobin intron 2, ODC introns 6 and 7 along with intervening exon 7, and β-fibrinogen intron 5), together encompassing 2072 aligned positions, to infer the relationships within the African clade. The resulting tree is generally well supported and indicates that none of the three largest currently recognized genera are monophyletic. For instance, the species included in Andropadus represent three different clades that are not each other's closest relatives. The montane species currently placed in that genus form a strongly supported clade, which is sister to Ixonotus , Thescelocichla, Baeopogon and Chlorocichla , although within this clade the genus Chlorocichla is polyphyletic. The remaining Andropadus species fall into two groups, one of these with A . importunus and A . gracilirostris , which along with Calyptocichla serina form a basal branch in the African greenbul radiation. In support of some previous studies the Leaf-love ( Pyrrhurus scandens ) is placed within Phyllastrephus . We also propose a new classification that reflects the phylogenetic relationships among African greenbuls.     

中国南蛇藤属(卫矛科)一新记录种及其意义

在开展南蛇藤属分类修订的过程中,于2011年在中国云南省西双版纳傣族自治州勐腊县发现南蛇藤属中国分布新记录——拟独子藤（Celastrusmonospermoides）。该种形态与独子藤（C．monospermus）较为相似,主要区别为前者叶片窄椭圆形,先端渐尖,果实成熟后不具雌蕊柄,种子长12mm以内;后者叶片椭圆形、阔椭圆形至倒卵椭圆形,果实具有明显的雌蕊柄,种子长15Inn以上。分子系统学研究表明,拟独子藤与独子藤处于不同分支中,与青江藤（C．hindsii）系统关系最近。由于原记录仅分布于印度尼西亚、马来西亚和菲律宾等热带地区,因此在原产地和我国云南省之间的地区应有更多居群分布,这为探讨南蛇藤属的起源和扩散提供了线索。     

Molecular evidence for an Asian origin and a unique westward migration of species in the genus Castanea via Europe to North America

The genus Castanea (Fagaceae) is widely distributed in the deciduous forests of the Northern Hemisphere. The striking similarity between the floras of eastern Asia and those of eastern North America and the difference in chestnut blight resistance among species has been of interest to botanists for a century. To infer the biogeographical history of the genus, the phylogeny of Castanea was estimated using DNA sequence data from different regions of the chloroplast genome. Sequencing results support the genus Castanea as a monophyletic group with Castanea crenata as basal. The three Chinese species form a strongly supported sister clade to the North American and European clade. A unique westward expansion of extant Castanea species is hypothesized with Castanea originating in eastern Asia, an initial diversification within Asia during the Eocene followed by intercontinental dispersion and divergence between the Chinese and the European/North American species during the middle Eocene and a split between the European and the North American species in the late Eocene. The differentiation within North America and China might have occurred in early or late Miocene. The North America species are supported as a clade with C. pumila var. ozarkensis, the Ozark chinkapin, as the basal lineage, sister to the group comprising C. pumila var. pumila, the Allegheny chinkapin, and Castanea dentata, the American chestnut. Morphological evolution of one nut per bur in the genus may have occurred independently on two continents.