Intraspecific groups of Umbelopsis ramanniana inferred from nucleotide sequences of nuclear rDNA internal transcribed spacer regions and sporangiospore morphology

Umbelopsis ramanniana is a well-known species in this genus. A characteristic morphological feature of this fungus is the remarkable variation in the sporangiospore shape, which implies the genetic variations occur in the nucleotide sequences of the internal transcribed spacer (ITS) regions of the nuclear ribosomal DNA (nrDNA) in the U. ramanniana isolates. The relationship between the variations of the sequences of the nrDNA ITS regions and those of the sporangiospore morphology was investigated for 12 isolates of U. ramanniana collected in Europe. Neighbor-joining and parsimony analyses on the sequences suggested that these isolates split into three groups. Precise examination of the morphology showed that the isolates of those respective groups were different from each other in their sporangiospore shape. The present study implies at least three intraspecific groups exist in U. ramanniana and that the variations in the nucleotide sequences of the nrDNA ITS regions correlate well with those in the sporangiospore shape in these intraspecific groups.     

<Emphasis Type="Italic">Umbelopsis gibberispora</Emphasis> sp. nov. from Japanese leaf litter and a clarification of <Emphasis Type="Italic">Micromucor ramannianus </Emphasis>var. <Emphasis Type="Italic">angulisporus</Emphasis>

 Umbelopsis gibberispora is described as a new species in the genus Umbelopsis, Umbelopsidaceae, Mucorales. The species differs from others in this genus by ellipsoidal sporangiospores with unilaterally thickened walls. Phylogenetic analyses based on nuclear large subunit ribosomal DNA (nLSU rDNA) partial sequences suggest that U. gibberispora, U. swartii, and U. westeae form a clade together with the strains of Umbelopsis ramanniana. The ex-type strain of Micromucor ramannianus var. angulisporus is found to be very close to Umbelopsis vinacea, whereas other isolates identified under the former name in the sense of Linnemann fall in the U. ramanniana subclade. For these isolates, a new species, Umbelopsis angularis, is introduced. Phylogenetic relationships among Umbelopsis species are discussed related to their attributes of the sporangial wall and mature spore shapes. Received: August 27, 2002 / Accepted: March 11, 2003 Acknowledgments We thank Dr. Takashi Ohsono, Graduate School of Agriculture, Kyoto University, Japan, for providing the strain of U. gibberispora (CBS 109328). We also thank Dr. Wieland Meyer, University of Sydney, Australia for access to the phylogenetic tree based on ITS sequence data before publishing, and Dr. Richard C. Summerbell, Centraalbureau von Schimmelcultures, the Netherlands, for linguistic corrections.     

Intraspecific diversity and distribution of the cosmopolitan species Pseudo‐nitzschia pungens (Bacillariophyceae): morphology,genetics, and ecophysiology of the three clades

Three clades of Pseudo‐nitzschia pungens, determined by the internal transcribed space (ITS) region, are distributed throughout the world. We studied 15 P. pungens clones from various geographical locations and confirmed the existence of the three clades within P. pungens, based on ITS sequencing and described the three subgroups (IIIaa, IIIab, and IIIb) of clade III. Clade III (clade IIIaa) populations were reported for the first time in Korean coastal waters and the East China Sea. In morphometric analysis, we found the ultrastructural differences in the number of fibulae, striae, and poroids that separate the three clades. We carried out physiological tests on nine clones belonging to the three clades growing under various culture conditions. In temperature tests, only clade III clones could not grow at lower temperatures (10°C and 15°C), although clade I and II clones grew well. The estimated optimal growth range of clade I clones was wider than that of clades II and III. Clade II clones were considered to be adapted to lower temperatures and clade III to higher temperatures. In salinity tests, clade II and III clones did not grow well at a salinity of 40. Clade I clones were regarded as euryhaline and clade II and III clones were stenohaline. This supports the hypothesis that P. pungens clades have different ecophysiological characteristics based on their habitats. Our data show that physiological and morphological features are correlated with genetic intraspecific differentiation in P. pungens.     

Comprehensive phylogeny of the laughingthrushes and allies (Aves,Leiothrichidae) and a proposal for a revised taxonomy

DNA phylogenies have gradually shed light on the phylogenetic relationships of the large babbler group. We focus in this study on the family Leiothrichidae (laughingthrushes and “song babblers”), which represents the largest clade of babblers in terms of species diversity. Our phylogeny includes all genera and 82% of the recognized species, using mitochondrial and nuclear loci. The sister group to Leiothrichidae is composed of the Pellorneidae (“jungle babblers”) plus the genus Alcippe. Within Leiothrichidae, four strongly supported primary clades (A–D) are recovered. Clade A includes Grammatoptila, Laniellus and Cutia. Clade B includes a large group of laughingthrushes, all of them classified in Trochalopteron. In Clade C, the two laughingthrushes endemic to southern India, T. fairbanki and T. cachinnans, which have recently been proposed to be placed in the newly erected genus Montecincla, form a sister clade to the group comprising the “song babblers” (Lioptila, Leiothrix, Heterophasia, Minla, Liocichla, Actinodura, Chrysominla, Siva, and Sibia). Clade D includes the African babblers (Turdoides, Phyllanthus, Kupeornis), Asian relatives (Argya, Acanthoptila, Chatarrhaea) and all remaining laughingthrushes (Garrulax). The time estimates suggest that the early diversification of the Leiothrichidae occurred in the mid‐Miocene, a period that corresponds to the diversification of many passerine groups in Asia. A revised taxonomic classification of the family is proposed in the light of these results.     

Subfamilial relationships within Caryophyllaceae as inferred from 5' ndhF sequences

DNA sequences of the 5' end of the chloroplast ndhF gene for 15 species of Caryophyllaceae have been analyzed by parsimony and neighbor-joining analyses. Three major clades are identified, with little or no support for monophyly of traditionally recognized subfamilies. The first of the three major clades identified (Clade I) is constituted by part of the subfamily Paronychioideae. It includes members of the tribe Paronychieae and members of tribe Polycarpeae. The second (Clade II) contains members of the Paronychieae exclusively. Tribe Paronychieae is thus apparently polyphyletic and tribe Polycarpeae is at least paraphyletic. The third clade (Clade III) includes members of subfamilies Alsinoideae and Caryophylloideae along with the genus Spergularia. The genus Scleranthus is also part of Clade III, while Drymaria groups with the other genera of tribe Polycarpeae in Clade II. We conclude that morphological characters previously used to delimit subfamilial groupings in the Caryophyllaceae are apparently unreliable estimators of phylogeny.     

Morphological and phylogenetic analyses of <Emphasis Type="Italic">Uromyces appendiculatus</Emphasis> and <Emphasis Type="Italic">U. vignae</Emphasis> on legumes in Japan

Uromyces appendiculatus, inclusive of three varieties, is distinguished from U. vignae primarily by the position of urediniospore germ pores and putative host specificity. However, opinions concerning these morphological and physiological features as taxonomic characters have varied greatly, and distinction of these species has often been confused. To clarify the taxonomy of these two species, morphological features of urediniospores and teliospores of 225 rust fungus specimens on species of Phaseolus, Vigna, Apios, Lablab, and Dunbaria were examined by light microscopy and scanning electron microscopy. Forty-five specimens were subjected to molecular phylogenetic analyses. As a result, the position of germ pores in urediniospores and the teliospore-wall thickness were considered as good characters to separate three morphological groups. In molecular analyses, the specimens fell into two and three clades based on the nucleotide sequence at D1/D2 domain of LSU rDNA and ITS regions, respectively. One of the D1/D2 clades corresponded to one morphological group whereas another D1/D2 clade included two other morphological groups. In contrast, each of the three ITS clades corresponded to a separate morphological group. Neither morphological groups nor molecular clades were host limited. It is suggested that the three morphological groups that corresponded to three distinct ITS clades constitute distinct species.Contribution no. 186 from the Laboratory of Plant Parasitic Mycology, Institute of Agriculture and Forestry, University of Tsukuba, Japan     

Rhinomonas nottbecki n. sp. (Cryptomonadales) and Molecular Phylogeny of the Family Pyrenomonadaceae

The cryptomonad Rhinomonas nottbecki n. sp., isolated from the Baltic Sea, is described from live and fixed cells studied by light, scanning, and transmission electron microscopy together with sequences of the partial nucleus‐ and nucleomorph‐encoded 18S rRNA genes as well as the nucleus‐encoded ITS1, 5.8S, ITS2, and the 5′‐end of the 28S rRNA gene regions. The sequence analyses include comparison with 43 strains from the family Pyrenomonadaceae. Rhinomonas nottbecki cells are dorsoventrally flattened, obloid in shape; 10.0–17.2 μm long, 5.5–8.1 μm thick, and 4.4–8.8 μm wide. The inner periplast has roughly hexagonal plates. Rhinomonas nottbecki cells resemble those of Rhinomonas reticulata, but the nucleomorph 18S rRNA gene of R. nottbecki differs by 2% from that of R. reticulata, while the ITS region by 11%. The intraspecific variability in the ITS region of R. nottbecki is 5%. In addition, the predicted ITS2 secondary structures are different in R. nottbecki and R. reticulata. The family Pyrenomonadaceae includes three clades: Clade A, Clade B, and Clade C. All Rhinomonas sequences branched within the Clade C, while the genus Rhodomonas is paraphyletic. The analyses suggest that the genus Storeatula is an alternating morphotype of the genera Rhinomonas and Rhodomonas and that the family Pyrenomonadaceae includes some species that were described multiple times, as well as novel species.     

Phylogenetic Reappraisal of the Genus Monomorphina (Euglenophyceae) Based on Molecular and Morphological Data

A morphological and molecular examination of the genus Monomorphina was conducted on 46 strains isolated mainly from Korea. The strains were divided into two types based on morphological data: Monomorphina aenigmatica and M. pyrum ‐ like species. Phylogenetic analysis based on a combined data set of nuclear SSU and LSU and plastid SSU and LSU rDNA showed that the strains could be divided into eight clades: Clade A of M. aenigmatica, Clade B of the isolates (M. pyropsis) from Michigan, USA, Clade C of M. pseudopyrum, Clade D of the isolates (M. pyroria) from Bremen, Germany, Clade E of M. soropyrum, Clade F of M. pyriformis, Clade G of M. parapyrum, and Clade H of M. pyrum. Six of these clades came from strains that would be considered M. pyrum sensu Kosmala et Zakry?, one of which could be recognized as a traditional species (M. pyrum) and five were designated as new species; each species had unique molecular signatures at nr SSU rDNA helix 17 and 17′ and spacer E23_14′‐E23_15. The species of Monomorphina had a wide range of genetic diversity with interspecies sequence similarity of 85.6%–97.1% and intraspecies similarity of 96.4%–99.9%. Our results suggested that genetic diversity found in the M. pyrum complex justifies the recognition of a minimum of eight species within this genus, based on specific molecular signatures and gene divergence of the nr SSU rDNA sequences.     

PHYLOGENY OF PHOTOSYNTHETIC EUGLENOPHYTES BASED ON COMBINED CHLOROPLAST AND CYTOPLASMIC SSU RDNA SEQUENCE ANALYSIS1

Eighteen new 16S rDNA and 16 new 18S rDNA sequences from 24 strains, representing 23 species of photoautotrophic euglenoids, were obtained in nearly their entire length. Maximum parsimony, maximum likelihood, and Bayesian phylogenetic analyses were performed on separate data (39 sequences of 16S rDNA and 58 sequences of 18S rDNA), as well as on combined data sets (37 sequences). All methods of sequence analysis gave similar results in those cases in which the clades received substantial support. However, the combined data set produced several additional well‐supported clades, not encountered before in the analyses of green euglenoids. There are three main well‐defined clades (A, B/C/D, and G) on trees from the combined data set. Clade G diverges first, while clades A and B/C/D form sister groups. Clade A consists of Euglena species sensu stricto and is divided into three sub‐clades (A1, A2, and A3). Clade A3 (composed of E. deses and E. mutabilis) branches off first; then, two sister clades emerge: A1 (composed of E. viridis‐like species) and A2 (consisting of E. agilis and E. gracilis species). Clade B/C/D consists of the Strombomonas, Trachelomonas, Cryptoglena, Monomorphina, and Colacium genera. Clade G comprises Phacus and Lepocinclis, as well as the Discoglena species of Euglena, with Discoglena branching off first, and then Phacus and Lepocinclis emerging as sister groups.     

LOW MOLECULAR WEIGHT CARBOHYDRATES OF THE BANGIOPHYCIDAE (RHODOPHYTA)1

In the order Porphyridiales there are three clades based on molecular evidence. These show parallels with the low molecular weight carbohydrate (LMWCs) in different genera. Clade Porphyridiales 1 includes Dixoniella, Glaucosphaera, Rhodella, and one undescribed genus (3987) that all contain mannitol. Clade Porphyridiales 2 comprises taxa of the Stylonematales Rhodosorus and Stylonema species and contains digeneaside and sorbitol, whereas Chroodactylon has only sorbitol. In clade Porphyridiales 3 Flintiella, Porphyridium, and the undescribed genus (3797) all possess only floridoside. In the Erythropeltidales Rhodochaete contains floridoside and digeneaside, Erythrotrichia species contain only floridoside, Sahlingia subintegra has floridoside and traces of D‐floridoside, and Smithora has L‐isofloridoside plus floridoside. In the Compsopogonales Boldia and Compsopogon have only floridoside. Within these genera as presently circumscribed, the LMWCs appear to be a reliable character to supplement the usual cytological characters.     

Molecular evolution of <Emphasis Type="Italic">Agaricus</Emphasis> species based on ITS and LSU rDNA sequences

Phylogenetic analyses of 62 isolates of 42 Agaricus and related secotioid species (A. inapertus, Gyrophragmium dunalii and Longula texensis) were conducted based on sequence data of the internal transcribed spacers (ITS) and partial large subunit (LSU) of ribosomal DNA. Bayesian, maximum likelihood and maximum parsimony analyses were used to reveal evolutionary groups within the genus Agaricus, while molecular clock analyses were carried out to obtain more information about the divergence times during the evolution of Agaricus within the Basidiomycota. Six major distinct clades were found within the genus with varying levels of support. These monophyletic groups suggested interspecific relationships both confirming and challenging previous morphological sections in several cases. Our results show that most morphological features likely have evolved in apparently similar ways multiple times independently during evolution, and that the secotioid A. inapertus, Gyrophragmium dunalii and Longula texensis evolved from Agaricus species in Clade I. A new name Agaricus aridicola, and a new combination Agaricus texensis are suggested to replace the names Gyrophragmium dunalii and Longula texensis, respectively. Molecular clock estimates for minimal age of separation of the genus Agaricus from its closest relatives were 32.63 ± 8.06 and 15.45 ± 3.82 Ma, respectively, using calibrations based on other molecular clock studies on fungi and fossil data. However, Agaricus likely diverged much earlier (73.30 ± 18.12 Ma), as suggested by the estimate based on the most robust calibration.     

Phylogeny of the genus Pythium and description of new genera

Phylogeny of the genus Pythium is analyzed based on sequences of the large subunit ribosomal DNA D1/D2 region and cytochrome oxidase II gene region of Pythium isolates and comprehensive species of related taxa belonging to the Oomycetes. The phylogenetic trees show that the genus Pythium is a highly divergent group and divided into five well- or moderately supported monophyletic clades. Each clade is characterized by sporangial morphology such as globose, ovoid, elongated, or filamentous shapes. Based on phylogeny and morphology, the genus Pythium (s. str.) is emended, and four new genera, Ovatisporangium, Globisporangium, Elongisporangium, and Pilasporangium, are described and segregated from Pythium s. lato.     

A new peronosporomycete, <Emphasis Type="Italic">Halioticida noduliformans</Emphasis> gen. et sp. nov., isolated from white nodules in the abalone <Emphasis Type="Italic">Haliotis</Emphasis> spp. from Japan

Four strains belonging to the Peronosporomycetes (formerly Oomycetes) were isolated from white nodules found on the mantle of three species of abalone. In artificial seawater, the four isolates formed fragments such as in the genus Haliphthoros, but the protoplasm constriction was weaker, and fragments were longer, with smaller spaces between them, than those of Haliphthoros. The four strains form one or more discharge tubes from each zoosporangium. The four strains were similar, but not identical, to the genus Haliphthoros based on morphological characteristics. As a result, the four isolates were classified in a new genus and species, Halioticida noduliformans gen. et sp. nov. Phylogenetic analysis of the D1/D2 region of the large subunit ribosomal RNA gene (LSU rDNA) was performed, and the four isolates showed 100%–99.8% concordance. In the phylogenetic tree, the four isolates were not classified in the subclass Peronosporomycetidae, Saprolegniomycetidae, or Rhipidiomycetidae. However, the four isolates formed a new clade with genera Haliphthoros and Halocrusticida in Peronosporomycetes. Within this new clade, the four isolates, Haliphthoros spp. and Halocrusticida spp., were grouped in their respective independent subclades. These results showed that these were the new genus and species from the morphological characteristics.     

Long‐standing environmental conditions,geographic isolation and host–symbiont specificity influence the relative ecological dominance and genetic diversification of coral endosymbionts in the genus Symbiodinium

Aim This study examines the importance of geographic proximity, host life history and regional and local differences in environment (temperature and water clarity) in driving the ecological and evolutionary processes underpinning the global patterns of diversity and distribution of symbiotic dinoflagellates. By comparing and contrasting coral–algal symbioses from isolated regions with differing environmental conditions, we may assess the potential of coral communities to respond to significant changes in climate. Location Indian Ocean. Methods Community assemblages of obligate symbiotic invertebrates were sampled at numerous sites from two regions, the north‐eastern Indian Ocean (Andaman Sea, western Thailand) and the western Indian Ocean (Zanzibar, Tanzania). Molecular genetic methods, including denaturing gradient gel electrophoresis analysis of the ribosomal internal transcribed spacers, DNA sequencing and microsatellite genotyping, were used to characterize the ‘species’ diversity and evolutionary relationships of symbiotic dinoflagellates (genus Symbiodinium). Host–symbiont specificity, geographic isolation and local and regional environmental factors were evaluated in terms of their importance in governing the distribution and prevalence of certain symbiont taxa. Results Host‐generalist symbionts (C3u and D1‐4, formerly D1a now designated Symbiodinium trenchi) frequently occurred alone and sometimes together in hosts with horizontal modes of symbiont acquisition. However, the majority of Symbiodinium diversity consisted of apparently host‐specific ‘species’. Clade C Symbiodinium were diverse and dominated host assemblages from sites sampled in the western Indian Ocean, a pattern analogous to symbiont communities on the Great Barrier Reef with similar environmental conditions. Clade D Symbiodinium were diverse and occurred frequently in hosts from the north‐eastern Indian Ocean, especially at inshore locations, where temperatures are warmer, water turbidity is high and large tidal exchanges commonly expose coral populations to aerial desiccation. Main conclusions Regional and local differences in cnidarian–algal combinations indicate that these symbioses are ecologically and evolutionarily responsive and can thrive under various environmental conditions. The high temperatures and turbid conditions of the north‐eastern Indian Ocean partly explain the ecological success of Clade D Symbiodinium relative to Clade C. Phylogenetic, ecological and population genetic data further indicate that Clade D has undergone an adaptive radiation, especially in regions around Southeast Asia, during the Pleistocene.     

Chloroplast DNA phylogeography of <Emphasis Type="Italic">Pedicularis</Emphasis> ser. <Emphasis Type="Italic">Gloriosae</Emphasis> (Orobanchaceae) in Japan

Phylogeographic analyses using chloroplast DNA (cpDNA) variation were performed for Pedicularis ser. Gloriosae (Orobanchaceae). Eighty-one plants of 18 populations of 6 species (P. gloriosa, P. iwatensis, P. nipponica, P. ochiaiana, P. sceptrum-carolinum and P. grandiflora) were analyzed. Fifteen distinct haplotypes were identified based on six cpDNA regions: the intergenic spacer between the trnT and trnL 3′exon, trnL 3′exon-trnF, atpB-rbcL, accD–psaI, the rpl16 intron and the trnK region (including the matK gene). Via phylogenetic analyses of the haplotypes, two continental species, P. sceptrum-carolinum and P. grandiflora, were placed at the most ancestral position in the trees. The former species is widely distributed in the Eurasian continent, and the latter is distributed in Far East Asia. Two robust major cpDNA clades (clades I and II) were revealed in the Japanese archipelago, although the statistical values of monophyly of these clades were weak. Clade I included the haplotypes (A-1, A-2, B-1, B-2 and J) of three species (P. gloriosa, P. iwatensis and P. ochiaiana), and Clade II included seven haplotypes (C-D, E-1, E-2 and F-H) of P. nipponica. These results suggest that this series originated on the Eurasian continent and that subsequently populations at the eastern edge of the continent differentiated into the two Japanese lineages. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The Gray Toad-headed Agama,Phrynocephalus scutellatus,on the Iranian Plateau: The degree of niche overlap depends on the phylogenetic distance

The Gray Toad-headed Agama (Phrynocephalus scutellatus) occurs in Iran, Pakistan and Afghanistan and is represented in Iran by four distinctive genetic clades. We built distribution models for three of these clades (one clade was not included due to a low number of distribution records) using Maximum Entropy Algorithm in order to determine the contribution of ecological factors to the distribution pattern. The degree of spatial niche overlap between every pair of clades were measured using Schoener's D niche overlap metric. The results showed that at species-level climate variables (annual precipitation, annual mean temperature) were the most influential parameters determining the boundaries of the distribution in Iran. Temperature seasonality was found to be the most influential factor in the distribution of both Clade I and Clade II. However, this variable was replaced by the annual mean temperature for Clade VI. Based on the results of Schoener's D metric, Clades I and II had the lowest, and Clades II and VI the highest level of ecological niche overlap. Comparing the result of niche overlap with genetic distance between the clades, it was found that the ecologically least similar clades were those with the longer history of genetic segregation.     

Phylogenetic Analysis of Uromyces Species Infecting Grain and Forage Legumes by Sequence analysis of Nuclear Ribosomal Internal Transcribed Spacer Region

DNA sequence analysis of the nuclear ribosomal internal transcribed spacer region (ITS) was performed to determine phylogenetic relationship between 49 isolates of rusts infecting grain and forage legumes. Isolates were collected from different hosts and distinct geographic origins and represent eight species of Uromyces: U. anthyllidis, U. appendiculatus, U. ciceris‐arietini, U. minor, U. pisi, U. striatus, U. viciae‐fabae and U. vignae. ITS sequences revealed length polymorphisms and variation in DNA sequence that were used to characterize phylogenetic relationships by maximum parsimony, maximum likelihood and Bayesian analyses which in general agreed revealing the presence of four clearly distinct clades. Clade one included the isolates causing rust on chickpea, fenugreek and alfalfa. Clade two was composed by rust isolates of field clover and pea plants, while the third clade was formed by bean and cowpea isolates. Clade four was the largest and included all the rust isolates infecting faba bean. Within this clade, the highly supported subclusters of U. viciae‐fabae collected on Lens culinaris, U. viciae‐fabae collected on Vicia sativa and U. viciae‐fabae collected on Lathyrus palustris suggest an ongoing process of host specialization.     

Bradyrhizobium canariense and Bradyrhizobium japonicum are the two dominant rhizobium species in root nodules of lupin and serradella plants growing in Europe

Forty three Bradyrhizobium strains isolated in Poland from root nodules of lupin species (Lupinus albus, L. angustifolius and L. luteus), and pink serradella (Ornithopus sativus) were examined based on phylogenetic analyses of three housekeeping (atpD, glnII and recA) and nodulation (nodA) gene sequences. Additionally, seven strains originating from root-nodules of yellow serradella (O. compressus) from Asinara Island (Italy) were included in this study. Phylogenetic trees revealed that 15 serradella strains, including all yellow serradella isolates, and six lupin strains grouped in Bradyrhizobium canariense (BC) clade, whereas eight strains from pink serradella and 15 lupin strains were assigned to Bradyrhizobium japonicum (BJ1). Apparently, these species are the two dominant groups in soils of central Europe, in the nodules of lupin and serradella plants. Only three strains belonged to other chromosomal lineages: one formed a cluster that was sister to B. canariense, one strain grouped outside the branch formed by B. japonicum super-group, and one strain occupied a distant position in the genus Bradyrhizobium, clustering with strains of the Rhodopseudomonas genus. All strains in nodulation nodA gene tree grouped in a cluster referred to as Clade II, which is in line with earlier data on this clade dominance among Bradyrhizobium strains in Europe. The nodA tree revealed four well-supported subgroups within Clade II (II.1-II.4). Interestingly, all B. canariense strains clustered in subgroup II.1 whereas B. japonicum strains dominated subgroups II.2-II.4.