Systematics of green algae resembling Ulva conglobata,with a description of Ulva adhaerens sp. nov. (Ulvales,Ulvophyceae)

A phylogenetic and morphological study of green algae resembling Ulva conglobata from Japan was undertaken, along with morphological observations of the original material of U. conglobata Kjellman. The samples resembling U. conglobata included five genetically distinct species: U. fasciata, U. pertusa, U. tanneri, Ulva sp. 1 and Ulva sp. 2. The discovery of marginal denticulations in some of the original material of U. conglobata, made it possible to distinguish those species without denticulations: U. pertusa, U. tanneri and Ulva sp. 2. The morphological characteristics of Ulva sp. 1 matched those of U. conglobata, but Ulva sp. 1 was not clearly identified as U. conglobata owing to the lack of DNA sequence data of the original material. Ulva sp. 2 had lobes adhering to each other by rhizoids. This morphological feature is stable in Ulva sp. 2 and unique among Ulva species. In conjunction with the molecular data, Ulva sp. 2 was described as a new species, U. adhaerens sp. nov. This species features rhizoidal extensions in regions other than the base and an elaborate arrangement of the extensions used for adhesion. It thereby expands our knowledge of the morphogenesis of the morphologically simple genus Ulva.     

New species of freshwater Ulva, Ulva limnetica (Ulvales, Ulvophyceae) from the Ryukyu Islands, Japan

Ulva limnetica Ichihara et Shimada, sp. nov. (Ulvales, Ulvophyceae) is described from the Ryukyu Islands, Japan, and is characterized by thalli that are: (i) branched, tubular, fragile and wrinkled; (ii) up to 80 cm in height and up to 2 cm in diameter; (iii) light to yellowish green in color; and (iv) having an asexual reproduction by means of quadriflagellate swarmers. Rhizoidal cells bear tubular extensions on the outside of the cell layer in the stipe. Ulva limnetica is distinguished from species with similar thalli by chloroplast disposition, branching pattern, number of pyrenoids per cell and gross morphology. It is also distinguished by sequences of the nuclear-encoded 18S ribosomal RNA gene, internal transcribed spacer 2 region and the plastid-encoded large subunit of ribulose-1,5-bisphosphate carboxylase/oxgenase gene ( rbc L). Ulva limnetica was clustered with other Ulva species in an early diverging lineage within the genus.     

Confirmation of cultivated Porphyra tenera (Bangiales, Rhodophyta) by polymerase chain reaction restriction fragment length polymorphism analyses of the plastid and nuclear DNA

Polymerase chain reaction restriction fragment length polymorphism (PCR‐RFLP) analysis of the plastid ribulose‐1,5‐bisphosphate carboxylase (RuBisCo) spacer region was developed for a more reliable and rapid species identification of cultivated Porphyra in combination with PCR‐RFLP analysis of the nuclear internal transcribed spacer (ITS) region. From the PCR‐RFLP analyses of the plastid and nuclear DNA, we examined seven strains of conchocelis that were used for cultivation as Porphyra tenera Kjellman but without strict species identification. The PCR‐RFLP analyses suggested that two strains, C‐32 and 90‐02, were cultivated P. tenera and that the other five strains, C‐24, C‐28, C‐29, C‐30 and M‐1, were Porphyra yezoensis f. narawaensis Miura. To identify species more accurately and to reveal additional genetic variation, the two strains C‐32 and 90‐02 were further studied by sequencing their RuBisCo spacer and ITS‐1 regions. Although RuBisCo spacer sequences of the two strains were identical to each other, each of their ITS‐1 sequences showed a single substitution. The sequence data again confirmed that the two strains (C‐32 and 90‐02) were cultivated P. tenera, and suggested that the two strains showed some genetic variation. We concluded that PCR‐RFLP analysis of the plastid and nuclear DNA is a powerful tool for reliable and rapid species identification of many strains of cultivated Porphyra in Japan and for the collection of genetically variable breeding material of Porphyra.     

Phylogeographical structure, distribution and genetic variation of the green algae Ulva intestinalis and U. compressa (Chlorophyta) in the Baltic Sea area

The marine algae Ulva intestinalis and U. compressa are morphologically plastic with many overlapping characters and are therefore difficult to distinguish from each other. The present distribution of U. intestinalis and U. compressa is investigated along the salinity gradient in the Baltic Sea area through analyses of internal transcribed spacer (ITS) sequence data. Also, the amount and distribution of intraspecific genetic polymorphism in the ITS region is studied allowing inferences on the phylogeographical pattern and postglacial recolonization of the Baltic Sea area. The data show that of the two species only U. intestinalis occurs in the Baltic Sea. The distribution of U. compressa is more restricted than previously reported, and it was not found in salinities lower than 15 ppt. All of Scandinavia and the Baltic Sea were covered with ice during the last ice age and the organisms in the Baltic Sea must have colonized the area after the ice had started to melt. The genetic diversity of U. intestinalis and U. compressa in the Baltic Sea and the neighbouring area was found to be reduced compared to that in the British Isles. This reduction may be the result of either a historical reduction of diversity or an adaptation of specific clones to the northern environmental conditions.     

PHYLOGENETIC RELATIONSHIPS WITHIN THE COLONIAL VOLVOCALES (CHLOROPHYTA) INFERRED FROM rbcL GENE SEQUENCE DATA

The chloroplast-encoded large subunit of the ribulose-1, 5-bisphosphate carboxylase / oxygenase (rbcL) gene was sequenced from 20 species of the colonial Volvocales (the Volvacaceae, Goniaceae, and Tetrabaenaceae) in order to elucidate phylogenetic relationships within the colonial Volvocales. Eleven hundred twenty-eight base pairs in the coding regions of the (rbcL) gene were analyzed by the neighbor-joining (NJ) method using three kinds of distance estimations, as well as by the maximum parsimony (MP) method. A large group comprising all the anisogamous and oogamous volvocacean species was resolved in the MP tree as well as in the NJ trees based on overall and synonymous substitutions. In all the trees constructed, Basichlamys and Tetrabaena (Tetrabaenaceae) constituted a very robust phylogenetic group. Although not supported by high bootstrap values, the MP tree and the NJ tree based on nonsynonymous substitutions indicated that the Tetrabaenaceae is the sister group to the large group comprising the Volvocaceae and the Goniaceae. In addition, the present analysis strongly suggested that Pandorina and Astrephomene are monophyletic genera whereas Eudorina is nonmonophyletic. These results are essentially consistent with the results of the recent cladistic analyses of morphological data. However, the monophyly of the Volvocaceae previously supported by four morphological synapomorphies is found only in the NJ tree based on nonsynonymous substitutions (with very low bootstrap values). The genus Volvox was clearly resolved as a polyphyletic group with V. rousseletii Pocock separated from other species of Volvox in the rbcL gene comparisons, although this genus represents a monophyletic group in the previous morphological analyses. Furthermore, none of the rbcL gene trees supported the monophyly of the Goniaceae; Astrephomene was placed in various phylogenetic positions .     

Tribal delimitation and phylogenetic relationships of Loteae and Coronilleae (Faboideae: Fabaceae) with special reference to Lotus: evidence from nuclear ribosomal ITS sequences

The temperate herbaceous tribes Loteae and Coronilleae have traditionally been regarded as taxonomically distinct entities. More recent morphological assessments, however, have challenged this view and suggest combining the two tribes under Loteae. Two key features used to distinguish the Coronilleae from Loteae include jointed fruits and branched root nodules. We evaluate the taxonomic utility of these characters using information derived from phylogenetic analyses of the internal transcribed spacers ITS1 + 2, and the intervening 5.8S region of nuclear ribosomal DNA. Results from this study show that neither the Loteae nor Coronilleae form individual monophyletic groups, and that key fruit and root nodule characters used to distinguish the Coronilleae are homoplastic. Given these data, we support the recognition of a single tribe, Loteae. We also find that Lotus, the largest and most morphologically complex genus in either tribe, is not monophyletic. Rather, it consists of two geographically distinct lineages, Old and New World, each of which are more closely related to other Loteae genera: Old World Lotus are more closely related to Old World Anthyllis, while New World Lotus show closer affinities to Old World Coronilla. These data also have important implications for the biogeography of New World Lotus: equally most parsimonious reconstructions suggest a complex scenario of intercontinental dispersals that involve not only Old World Lotus but Coronilla as well.     

Molecular relationships in Encephalartos (Zamiaceae, Cycadales) based on nucleotide sequences of nuclear ITS 1&2, rbcL, and genomic ISSR fingerprinting

The cycad genus Encephalartos is restricted to Africa and is threatened with extinction in most of its range. Total DNA was extracted from 51, i.e., 78 %, of the described species of Encephalartos. The accessions were sampled from the furthest western occurrence of the genus in Nigeria, via Sudan and Uganda, to southern South Africa. The sequences of nuclear ribosomal internal transcribed spacer regions 1 and 2 (ITS 1&2), the chloroplast encoded rbcL gene, and ISSR genomic fingerprinting were employed to resolve the molecular history and the relationships within the genus. Sequence alignment, as well as ISSR fingerprinting, data show low genetic variation among all analysed accessions, indicating diversification within the Pliocene/Pleistocene. ITS 1&2 data agree well with morphological and geographical characters and resolved three major genetic clusters with overlapping distribution ranges in eastern South Africa. This area, that contains the largest diversity of genotypes of Encephalartos, may have served as a Pliocene/Pleistocene refugium.     

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.     

Evidence for the Polyphyly of Haworthia (Asphodelaceae Subfamily Alooideae; Asparagales) Inferred from Nucleotide Sequences of rbcL, matK, ITS1 and Genomic Fingerprinting with ISSR-PCR

Abstract: Four molecular markers have been studied to examine the phylogenetic position of the South African plant genus Haworthia Duval within the succulent Asphodelaceae. Sequence data of the chloroplast genes mat K and rbc L were compared to the nuclear markers ITS1 and ISSR (Inter Simple Sequence Repeat) analysis. Both lines of molecular data, chloroplast and nuclear DNA, indicate that Haworthia is polyphyletic, forming two distinct clades. Most taxa previously combined as Haworthia subgenus Haworthia branch off early in the alooid chloroplast trees forming a strongly monophyletic group, whereas subgenus Hexangulares forms a polyphyletic assemblage comprising other alooid genera. The nuclear markers ITS1 and ISSR fingerprinting support the two groups as distinctly different, therefore confirming the division seen in chloroplast DNA. The practical implication is that the generic concept of Haworthia may have to be restricted to H. subgenus Haworthia or alternatively, that the groups of Haworthia be treated as infrageneric taxa within a broadened (Linnaean) concept of Aloe.     

Systematics of Dendrobiinae (Orchidaceae), with special reference to Australian taxa

The systematics of subtribe Dendrobiinae, Dendrobium, and Australian dendrobiums in particular, are reviewed. Molecular approaches have delimited the subtribe, established Asian and Australasian clades and clarified relationships at some levels within them. At most lower taxonomic levels within Dendrobium s.l., resolution has been partial. Morphological and molecular phylogenetics indicate that Dendrocoryne, Rhizobium and some other sections are not monophyletic, with morphological analyses providing the greatest resolution into groups corresponding with biogeography. For most sections, sampling is too incomplete to determine relationships and monophyly. Distribution, pollination syndromes, breeding systems, fertility and specialized anatomical features contribute little to classification. Natural hybrids are more common than previously stated and may have played a role in the high level of homoplasy and evolution of Australian epiphytes. There is little support for splitting Dendrobium into many genera and species based on terminal clades. The evidence is interpreted as favouring recombination of Cadetia, Flickingeria and Diplocaulobium within the conserved broad genus Dendrobium. Nomenclatural stability is retained in continuing to use the long‐held subgeneric sectional classification, including many of the sections of Schlechter, and infraspecific ranks of subspecies and varieties. In Dendrocoryne, Thelychiton, Tropilis and Tetrabaculum are rejected, as is Dockrillia in Rhizobium. Proposals to elevate infraspecific taxa to species level for variable species complexes are not supported by current morphological or molecular data based on slowly evolving DNA regions. © 2011 The Linnean Society of London, Botanical Journal of the Linnean Society, 2011, 166 , 105–126.     

Molecular assessment of Ulva (Ulvales,Chlorophyta) diversity in Vietnam including the new species U. vietnamensis

Species diversity of Ulva in Vietnam was investigated using three commonly used genetic markers, the nuclear encoded rDNA ITS region and the plastid encoded rbcL and tufA genes. Single locus species delimitation methods, complemented with morphological and ecological information resulted in the delimitation of 19 species. This diversity is largely incongruent with the traditional understanding of Ulva diversity in Vietnam. Only four species identified in this study, U. lactuca, U. reticulata, U. spinulosa, and U. flexuosa, have been previously reported, and seven species, U. ohnoi, U. tepida, U. chaugulii, U. kraftiorum, U. meridionalis, U. limnetica, and U. aragoënsis, are recorded for the first time from Vietnam. Seven genetic clusters could not be associated with species names with certainty. A new species, U. vietnamensis, is described from marine to brackish coastal areas from southern Vietnam based on its morphological and molecular distinctiveness from the currently known Ulva species. A comparison with recent molecular-based studies of Ulva diversity showed that species composition in Vietnam is similar to that of adjacent countries, including Japan, China, as well as Australia. Our study emphasizes the importance of molecular data in the assessment of Ulva diversity, and indicates that a lot of diversity may still remain to be discovered, especially in tropical regions.     

碳源种类及碳氮比对眼点拟微绿球藻生长密度、油脂含量和脂肪酸组成的影响

研究了三种碳源Na2CO3、NaHCO3、葡萄糖对眼点拟微绿球藻生长密度和油旨含量的影响,实验结果表明相对于葡萄糖,无机碳源NaHCO3更利于眼点拟微绿球藻的生长.以NaHCO3为碳源,研究了在不同的接种密度、NaNO3浓度下,C/N对眼点拟微绿球藻生长密度和油脂含量的影响.实验结果表明,C/N对眼点拟微绿球藻生长密度的影响与接种密度和NaNO3浓度有关,在高的NaNO3浓度时,C/N对眼点拟微绿球藻生长密度的影响很小;在低的NaNO3浓度时,随着C/N比的增加,微绿球藻的生长密度先增加后下降,存在最佳的C/N比.最佳的C/N比随接种密度而变化,在接种密度为OD440=0.10时,最佳C/N比为3,当接种密度提高到OD440=0.70时,最佳C/N比增加到5.NaNO3浓度和C/N对微藻油脂含量均有较大影响,在不同的接种密度和NaNO3浓度下都表现为C/N=1时最利于微藻油脂的积累,这与卡尔文循环过程中核酮糖-1,5-二磷酸羧化酶/加氧酶的活性有关.本实验的最佳产油培养条件为以NaHCO3为碳源,初始接种密度为OD440=0.70,C/N=1∶1,CNaNO3=0.225g/L,此时油脂产率为56.7 mg/(L·d),EPA产率为6.5 mg/(L·d).     

Role of pyrenoids in the CO2-concentrating mechanism: comparative morphology, physiology and molecular phylogenetic analysis of closely related strains of Chlamydomonas and Chloromonas (Volvocales)

The morphology of the pyrenoid and the physiology of the CO₂-concentrating mechanism (CCM) were investigated in Chlamydomonas (Cd.) mutabilis Gerloff UTEX 578, Cd. radiata Deason et Bold UTEX 966, Cd. augustae Skuja UTEX 1969, Cd. macrostellata Lund SAG 72.81, Cd. bipapillata Bourrelly SAG 11-47, and Chloromonas (Cr.) insignis Gerloff et Ettl NIES-447, all of which are closely related phylogenetically to the pyrenoid-less strains of Chloromonas. In the chloroplasts of Cd. mutabilis UTEX 578, Cd. radiata UTEX 966, Cd. augustae UTEX 1969, and Cd. macrostellata SAG 72.81, a typical, spheroidal, electron-dense pyrenoid matrix surrounded by starch granules was present, and ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco; EC 4.1.1.39) molecules were highly concentrated in the pyrenoid matrix. On the other hand, while the pyrenoid matrix of Cr. insignis NIES-447 was electron-dense that of Cd. bipapillata SAG 11-47 was not, and neither was surrounded by starch granules. The pyrenoid matrices of these two species exhibited a higher concentration of Rubisco molecules than the thylakoid region (thylakoid and stroma) of the chloroplasts; however, the densities of Rubisco molecules in these pyrenoid matrices were low compared with those of the other four Chlamydomonas strains examined in this study and that of Cd. reinhardtii Dangeard. In all six strains examined, the presence of the CCM was indicated by relatively high photosynthetic affinities for CO₂ (low values of K_0.5(CO₂)). However, differences in the inorganic carbon (Ci) pools were recognized in relation to the differences in pyrenoid morphology among the strains. In the typical pyrenoid-containing strains. Cd. mutabilis UTEX 578 and Cd. radiata UTEX 966, the ratio of internal to external inorganic carbon was about 20, while in Cr. insignis NIES-447 and Cd. bipapillata SAG 11-47 the ratio was only 2–3 similar to the two pyrenoid-less, CCM-containing strains of Chloromonas previously examined (E. Morita et al., 1998, Planta 204: 269–276). It is thus speculated that the presence of typical pyrenoids with a high concentration of Rubisco molecules is related to the formation of large Ci pools in the CCM. Detailed phylogenetic relationships among these Chlamydomonas/Chloromonas strains and the pyrenoid-less Chloromonas strains previously investigated were inferred based on the sequence of rbcL, the gene for the large subunit of Rubisco. Two monophyletic groups were resolved with high bootstrap values. Based on the tree topology resolved, it was inferred that loss of the typical pyrenoids accompanied by a decrease in intracellular Ci pools might have taken place independently in the two groups. Received: 21 August 1998 / Accepted: 30 November 1998     

Towards a monophyletic genus Albinaria (Gastropoda, Pulmonata): the first molecular study into the phylogenetic position of eastern Albinaria species

Recent molecular phylogenetic studies of the terrestrial snail genus Albinaria have caused a radical reassessment of its taxonomy. These studies, however, were limited to western species. This paper examines mitochondrial 12S sequences and nuclear ITS1 & 2 sequences of both eastern and western species, and demonstrates that Albinaria, in its most recent definition, is not monophyletic. Both molecular datasets place ' Albinaria ' hedenborgi from Lebanon in a well-supported clade with species of the genus Cristataria , distributed south-east of the vicariant range of Albinaria . The remaining species from Cyprus, Turkey and Greece constitute a well-supported monophyletic group. These two clades form geographical clusters, whereas Albinaria in the current definition (including ' A. '  hedenborgi ) has a disjunct distribution. ' A.' hedenborgi should therefore be classified with Cristataria , together with the morphologically similar and geographically close ' A. '  nadimi .  © 2005 The Linnean Society of London, Zoological Journal of the Linnean Society , 2005, 143 , 531−542.     

Molecular,biochemical and morphological data suggest an affiliation of Spongiochrysis hawaiiensis with the Trentepohliales (Ulvophyceae,Chlorophyta)

The aeroterrestrial, unicellular green alga Spongiochrysis hawaiiensis had been included in the ulvophycean order Cladophorales based on small subunit (SSU) rDNA sequence data, and represents so far the only fully terrestrial member of this order. Other characteristics of S. hawaiiensis that are atypical for Cladophorales include the presence of large amounts of carotenoids and a budding‐like mode of cell division. As the position of this terrestrial, unicellular alga in an order of aquatic, multicellular green algae is unusual, we re‐evaluated the phylogenetic relationships of this enigmatic organism based on supplementary SSU rDNA sequences as well as novel large ribosomal subunit (LSU) rDNA and internal transcribed spacer (ITS rDNA) sequences. Additionally, we examined several morphological characters of S. hawaiiensis, as well as low molecular weight carbohydrate (LMWC) patterns of S. hawaiiensis and members of the Cladophorales and Trentepohliales as potential chemotaxonomic markers. We found S. hawaiiensis to be uninucleate. The analysis of the LMWC content detected the presence of the polyol erythritol in S. hawaiiensis and in the Trentepohliales, while this compound was missing in the Cladophorales. The phylogenetic analyses of the novel sequences placed S. hawaiiensis in the terrestrial Trentepohliales. This placement is supported by the aeroterrestrial habitat, the presence of large amounts of carotenoids, the uninucleate cells, and the presence of the polyol erythritol as a protective compound against water loss.     

Studies on the fine structure of Glaucocystis nostochinearum Itzigs. II. Membrane morphology and taxonomy

The fine structure of the alga Glaucocystis nostochinearum Itzigs. has been studied by thinsectioning and freeze-etching. Many of the features already established by sectioning have been verified but different conclusions are drawn, particularly with regard to the photosynthetic organs. The action of glycerol solutions on the morphology of the plasmalemma as revealed by freeze-etching is shown to be of general importance for the freeze-etching of plant cells and of particular importance in elucidating the true form of the plasmalemma in Glaucocystis. The literature regarding the taxonomic position of Glaucocystis has been reviewed, and, along with a completely new demonstration of plasmalemma structure, an attempt has been made to reclassify this organism.     

Molecular systematics of Apiaceae subfamily Apioideae: phylogenetic analyses of nuclear ribosomal DNA internal transcribed spacer and plastid RPO C1 intron sequences

Evolutionary relationships among representatives of Apiaceae (Umbelliferae) subfamily Apioideae have been inferred from phylogenetic analyses of nuclear ribosomal DNA internal transcribed spacer (ITS 1 and ITS 2) and plastid rpoC1 intron sequences. High levels of nucleotide sequence variation preclude the use of the ITS region for examining relationships across subfamilial boundaries in Apiaceae, whereas the rpoC1 intron is more suitably conserved for family-wide phylogenetic study but is too conserved for examining relationships among closely related taxa. In total, 126 ITS sequences from subfamily Apioideae and 100 rpoC1 intron sequences from Apiaceae (all three subfamilies) and outgroups Araliaceae and Pittosporaceae were examined. Phylogenies estimated using parsimony, neighbor-joining, and maximum likelihood methods reveal that: (1) Apiaceae subfamily Apioideae is monophyletic and is sister group to Apiaceae subfamily Saniculoideae; (2) Apiaceae subfamily Hydrocotyloideae is not monophyletic, with some members strongly allied to Araliaceae and others to Apioideae + Saniculoideae; and (3) Apiaceae subfamily Apioideae comprises several well-supported subclades, but none of these coincide with previously recognized tribal divisions based largely on morphological and anatomical characters of the fruit. Four major clades in Apioideae are provisionally recognized and provide the framework for future lower level phylogenetic analyses. A putative secondary structure model of the Daucus carota (carrot) rpoC1 group II intron is presented. Of its six major structural domains, domains II and III are the most, and domains V and VI the least, variable.