Phylogeny of lobose amoebae based on actin and small-subunit ribosomal RNA genes

Lobose amoebae are abundant free-living protists and important pathogenic agents, yet their evolutionary history and position in the universal tree of life are poorly known. Molecular data for lobose amoebae are limited to a few species, and all phylogenetic studies published so far lacked representatives of many of their taxonomic groups. Here we analyze actin and small-subunit ribosomal RNA (SSU rRNA) gene sequences of a broad taxon sampling of naked, lobose amoebae. Our results support the existence of a monophyletic Amoebozoa clade, which comprises all lobose amoebae examined so far, the amitochondriate pelobionts and entamoebids, and the slime molds. Both actin and SSU rRNA phylogenies distinguish two well-defined clades of amoebae, the "Gymnamoebia sensu stricto" and the Archamoebae (pelobionts + entamoebids), and one weakly supported and ill-resolved group comprising some naked, lobose amoebae and the Mycetozoa.     

A preliminary investigation of the order Bangiales (Bangiophycidae, Rhodophyta) based on sequences of nuclear small-subunit ribosomal RNA genes

We investigated phylogenetic relationships among red algae of the order Bangiales by analysis of sequences of the nuclear gene encoding cytosolic small-subunit ribosomal RNA in Bangia atropurpurea (Roth) C. Ag. and eight samples representing seven species of Porphyra. The ssu-rDNA range from 1818 to 1845 nucleotides in length, with guanosine plus cytosine ratios between 47.0% and 48.6%. A group IC1 intron occurs in the B atropurpurea ssu-rDNAs at the same position as in P. spiralis var. amplifolia Oliveira Filho et Coll and several other eukaryote ssu-rDNAs. The nine sequences form a stable monophyletic group upon phylogenetic analysis. The ssu-rDNA from B. atropurpurea nests stably within the Porphyra group and is closely related to P. amplissima (Kjellm.) Setchell et Hus in Hus, making the genus Porphyra paraphyletic. No correlation is seen between phylogenetic position and number of cell layers in the Porphyra thallus. We discuss possible taxonomic and evolutionary implications of these observations.     

中国淡水鱼类寄生鞭毛虫两新种（动鞭毛纲：动体目）

本文报道寄生在泥鳅Misgurnus anguillicaudatus(Cantor)血液中的动鞭毛虫两新种,即泥鳅锥体虫Trypanosoms misgurni sp.nov.和重庆隐鞭虫Cryptobia chongqingensis sp.nov.。标本采自重庆市沙坪坝区和北碚区、模式标本保存于重庆师范学院生物学系和西南师范大学生物学系。     

Phylogenetic Relationships of Species of the Genus Kluyveromyces van der Walt (Saccharomycetaceae) Deduced from the Partial Sequences of 18S and 26S Ribosomal RNAs

In order to clarify the phylogenetic relationships of the species classified in the genus Kluyveromyces (Saccharomycetaceae), three partial base sequences of 18S and 26S rRNAs of eighteen strains were determined. The regions determined of the strains corresponded to positions 1451 through 1618 (168 bases) of 18S rRNA and to positions 1611 through 1835 (225 bases) and 493 through 622 (130 bases) of a strain (IFO 2376) of Saccharomyces cerevisiae. The analyses of the partial base sequences suggested that the genus Kluyveromyces is phylogenetically heterogeneous, ranging from the strains that are quite close to the strain of S. cerevisiae to the strains that are distinct enough to be classified in genera separate from the genus Saccharomyces. From our sequence data, we concluded that the extent of the genus Kluyveromyces should be restricted to only one species, K. polysporus, the type species of the genus. Kluyveromyces phaffii was also distinct enough to deserve another genus. Kluyveromyces cellobiovorus was not close to any of the strains of Kluyveromyces species examined, and should be excluded from the genus. Most of the strains of the species examined were fairly close to the strain of S. cerevisiae.     

由部分核糖体RNA序列确定的绿僵菌属种系统发育关系

本研究应用特异寡聚核苷酸引物和双脱氧核苷酸终止法测定绿僵菌Metarhizium不同种的部分rRNA序列。这些序列分别选自小亚基(18S)和大亚基(25S)rRNA上的不同区域。校排的不同序列用于计算核苷酸的差异。绿僵菌的系统发育关系采用最大可能性(Maximum-Likelihood)方法分析。所有的分类单元聚类成二个分支。在产生圆柱状瓶形小梗的种中,M.anisopliae var.anisopliae,M.anisopliae var.majus,M.brunneum(NRRL1944),M.guizhouense,M.pingshaense和未知种M.sp.2组成一个密切相关的群组,而M.anisopliae(ACCC 30104)位于这群组之外。在产生棍棒状瓶形小梗的种中,M.album,M.cylindrosporae和M.flavoviride互相独立成支。实验结果不支持Metarhizium只能区分为M.anisopliae和M.flavoviride二个种的分类观点。文中还讨论了经典分类标准的系统发育价值。     

Structural analyses of plastid-derived 16S rRNAs in holoparasitic angiosperms

Higher-order structures have been constructed for plastid-encoded small-subunit (SSU, 16S), rRNAs from representatives of seven nonphotosynthetic holoparasitic angiosperm families: Apodanthaceae, Cynomoriaceae, Cytinaceae, Balanophoraceae, Hydnoraceae, Mitrastemonaceae, and Rafflesiaceae. Whereas most pairwise comparisons among angiosperms differ by 2–3% in substitutions, the 16S rRNAs of the holoparasites show an increasingly greater number of mutations: Cynomorium (7.3%), Cytinus (8.0%), Bdallophyton (12.7%), Mitrastema (14.9%), Hydnora (19.4%), Pilostyles (30.4%) and Corynaea (35.9%). Despite this high level of sequence variation, SSU structures constructed for all species except Pilostyles possess the typical complement of 50 helices (that contain numerous compensatory mutations) thereby providing indirect evidence supporting their functionality. Pilostyles, likely with the most unusual plastid 16S rRNA yet documented, lacks four major helices and contains lengthy insertions for four others. Sequences of products generated via RT-PCR show that these structural modifications are present on a mature (transcribed) rRNA. The trend toward increasing numbers of base substitutions in the holoparasites is accompanied by a marked increase in AA+U content of the rRNA. This A/T drift phenomenon of rDNA is especially apparent in Corynaea whose SSU rDNA sequence is 72% A+T. A comparison of Cytinus to tobacco showed that substitution rates appear to be dependent upon the composition of neighboring bases. Transversions represented 26% of the mutations when flanking bases were G or C whereas transversions increased to 36% when the flanking bases were A to T. The underlying molecular mechanism associated with these high substitution rates is presently unknown, however, relaxation of selection pressure on ribosome function resulting in altered DNA replication and/or repair systems may be involved.     

Planktic foraminiferal molecular evolution and their polyphyletic origins from benthic taxa

Phylogenetic analyses based on partial sequences of the small subunit (SSU) ribosomal (r) RNA gene have shown that the planktic and benthic foraminifera form a distinct monophyletic group within the eukaryotes. In order to determine the evolutionary relationships between benthic and planktic foraminifers, representatives of spinose and non-spinose planktic genera have been placed within a molecular SSU rDNA phylogeny containing sequences of the benthic suborders available to date. Our phylogenetic analysis shows that the planktic foraminifers are polyphyletic in origin, not evolving solely from a single ‘globigerinid-like’ lineage in the Mid-Jurassic, but derived from at least two ancestral benthic lines. The benthic ancestor of Neogloboquadrina dutertrei may have entered the plankton later than the Mid-Jurassic, and further investigation of related extant species should provide an indication of the timing of this event. The evolutionary origin of the non-spinose species Globorotalia menardii remains unclear. The divergences of the planktic spinose species generally support recent phylogenies based on the fossil record, which infer a radiation from a globigerinid common ancestor in the Mid- to Late Oligocene. The branching pattern indicates that there are possibly four distinct groups within the main spinose clade, with large evolutionary distances being observed between them. Globigerinoides conglobatus clusters strongly with Globigerinoides ruber and are divergent from Globigerinella siphonifera, Orbulina universa and Globigerinoides sacculifer.Conserved regions of the SSU rRNA gene show sufficient variation to discriminate foraminifers at the species level. Large genetic differences have been observed between the pink and white forms of Gs. ruber and between Ge. siphonifera Type I and II. The two types of Ge. siphonifera cannot be discriminated by traditional palaeontological methods, which has considerable implications for tracing fossil lineages and for the estimation of molecular evolutionary rates based upon the fossil record. The conserved regions show a high degree of sequence identity within a species, providing signature sequences for species identification. The variable regions of the gene may prove informative for population level studies in some species although complete sequence identity was observed in G. sacculifer and O. universa between specimens collected from the Caribbean and Western Pacific.     

Phylogenetic Relationships Among Anuran Trypanosomes as Revealed by Riboprinting

Twenty trypanosome isolates from Anura (frogs and toads) assigned to several species were characterized by riboprinting–restriction enzyme digestion of polymerase chain reaction amplified small subunit ribosomal RNA genes. Restriction site polymorphisms allowed distinction of all the recognized species and no intraspecific variation in riboprint patterns was detected. Phylogenetic reconstruction using parsimony and distance estimates based on restriction fragment comigration showed Trypanosoma chattoni to be only distantly related to the other species, white T. ranarum and T. fallisi appear to be sister taxa despite showing non-overlapping host specificities.     

MOLECULAR SYSTEMATIC ANALYSES INDICATE THAT THE ENIGMATIC APOPHLAEA IS A MEMBER OF THE HILDENBRANDIALES (RHODOPHYTA, FLORIDEOPHYCIDAE)

Nuclear small-subunit (SSU) ribosomal DNA sequences were determined for Apophlaea lyallii Hooker f. et Harvey from New Zealand and an isolate of Hildenbrandia rubra (Sommerfelt) Meneghini from France. These data, in addition to SSU sequences representative of all the major florideophyte lineages, were used in molecular systematic investigations to confirm inclusion of Apophlaea in the Hildenbrandiales. Anatomical features that serve to unite Apophlaea and Hildenbrandia are discussed to buttress this result. Furthermore, our analyses support recognition of the Hildenbrandiales as sister to all other extant florideophyte lineages studied to date. An anomalous result positioning the SSU sequence generated herein for Hildenbrandia rubra closer to Apophlaea lyallii than to a previously published sequence for Hildenbrandia rubra requires further investigation.     

SMALL-SUBUNIT RIBOSOMAL DNA SEQUENCE ANALYSES AND A RECONSTRUCTION OF THE INFERRED PHYLOGENY AMONG SYMBIOTIC DINOFLAGELLATES (PYRROPHYTA)1

The small-subunit ribosomal RNA genes (SSU rDNA) from the four symbiotic dinoflagellates, Symbiodinium corculorum Trench isolated from the bivalve mollusc Corculum cardissa (from Belau, Western Caroline Is.), S. meandrinae Trench, from the scleractinian coral Meandrina meandrites (from famaica, W.I.), Gloeodinium viscum Banaszak et al. from the hydrocoral Millepora dichotoma (from the Gulf of Aqaba), and Amphidinium belauense Trench from the acoel flatworm Haplodiscus sp. (from Belau) have been amplified by the polymerase chain reaction, cloned, and sequenced. Following alignment of these complete sequences to homologous sequences from six other dinoflagellates, eight api-complexans, six ciliates, six chromophytes and oomycetes, three ascomycetes, two rhodophytes, two chlorophytes, and two myxomycetes (with Physarum polycephalum as the outgroup), phylogenetic reconstruction was conducted using Fitch and Margoliash distance, DNA maximum likelihood, and Wagner parsimony methods, with bootstrap resampling. All methods generated trees with similar topologies. The inferred “across Kingdom” phylogeny reemphasizes previous reports that show that the dinoflagellates, the apicomplexans, and the ciliates share a common ancestry and that the dinoflagellates are distantly related to the chromophyte-oömycete lineage. The evidence supports the concept of a polyphyletic origin of dinoflagellate-invertebrate symbioses, as symbiotic dinoflagellates represent seven genera in at least four orders. The three symbiotic species, S. corculorum, S. meandrinae, and S. pilosum, consistent with their morphological and biochemical similarities, cluster most closely. Symbiodinium pulchrorum Trench, the symbiontfrom the Hawaiian sea anemone Aiptasia pulchella, is more distantly related to them. Gloeodinium viscum is not closely related to the Symbiodinium species. Amphidinium carterae (free-living) and A. belauense (symbiotic) also appear to be distantly related to Symbiodinium. Some symbionts (e.g. S. corculorum, S. pilosum) from distant geographic locations (the Indo-Pacific and Caribbean, respectively) were found to be very closely related, whereas S. pulchrorum and S. corculorum from the Pacific were found to be distantly related. Analyses of 10 additional symbiotic and nonsymbiotic dinoflagellates, using partial SSU rDNA sequences to generate a tentative dinoflagellate phylogeny, indicate that members of the genus Symbiodinium cluster with most of the other (free-living) dinoflagellates in the genus Gymnodinium. The genus Amphidinium, as represented by A. carterae and A. belauense, appear to be distantly related to the other members of the Gymnodiniaceae. This analysis, combined with morphological and biochemical data, indicates that the symbionts S. pulchrorum (from Aiptasia pulchella) and S. bermudense Trench (from Aiptasia tagetes) from the Indo-Pacific and Caribbean, respectively, are very closely related but are not identical.     

小鲵科线粒体16S rRNA基因序列分析及其系统发育

To study the phylogeny of Hynobiidae, we amplified DNA fragments of 470 bp 16S ribosomal RNA (16S rRNA) gene on mitochondrial DNA from Ranodon sibiricus and Ranodon tsinpaensis. PCR products were cloned into PMD18 T vector after purification. These sequences were determined and deposited in the GenBank (accession numbers: AY373459 for Ranodon sibiricus, AY372534 for Ranodon tsinpaensis). By comparing the nucleotide differences of 16S ribosomal RNA sequences among Liua shihi, Pseudohynobius flavomaculatus and Batrachuperus genus from GenBank database, we analyzed the divergences and base substitution among these sequences with the MEGA software. The molecular results support that B. tibetanus, B. pinchonii and B. karlschmidti are classified into three valid species. Liua shihi has closer phylogenetic relationships to Ranodon tsinpaensis than to other species. More our results reveal that Pseudohynobius flavomaculatus is not a synonym of Ranodon tsinpaensis. [Acta Zoologica Sinica 50 (3) : 464 - 469,2004].     

Ultrastructural and phylogenetic studies on Blastocystis isolates from cockroaches

Four Blastocystis isolates from cockroaches were established and these isolates were morphologically confirmed as Blastocystis organisms by light and/or electron microscopy. As these isolates were morphologically indistinguishable from Blastocystis isolated from other animals, phylogenetic analyses were conducted using their small subunit ribosomal RNA genes. A analyses of these sequences with previously reported ones that had been classified into nine Blastocystis clades indicated the presence of a new clade that comprised only Blastocystis organisms from cockroaches (clade X). A clade comprised of amphibian and reptilian Blastocystis organisms (clade IX) was located at the basal position of the Blastocystis tree together with the common ancestor of Proteromonas and Protoopalina, clade X emerged after the divergences of these two basal clades and its branching position was clearly supported by bootstrap analysis.     

基于核糖体小亚基的波豆类鞭毛虫分子系统发育研究

波豆类鞭毛虫(动基体目)是一类非常重要的原生动物,结构独特,分布广泛.但是,这一类群的系统发育关系尚存有很多争议.为了更好的了解该类群的系统发育关系,作者分离、纯化并培养了Bodo designis DH,测定了它的SSU rRNA序列.根据该序列和GenBank中的相关序列,用最大简约法和邻接法分别构建了基于全序列和保守区序列的系统树.结果如下:1)波豆属是多系发育的;2)波豆类鞭毛虫和锥虫类鞭毛虫的系统发育关系仍有待于深入研究;3)虽然所得的系统树都显示出一种基本的双歧式树型结构,但采用不同方法所构建的系统树之间有明显的差异.这些似乎表明,就波豆类鞭毛虫而言,SSU rRNA不是一个令人满意的系统发育标记.     

A MOLECULAR ANALYSIS OF ANALIPUS AND RALFSIA (PHAEOPHYCEAE) SUGGESTS THE ORDER ECTOCARPALES IS POLYPHYLETIC1

We report partial 18S ribosomal DNA sequences of Analipus japonicus (Harvey) Wynne and Ralfsia fungiformis (Grunnerus) Setchell et Gardner. These sequences were compared with the corresponding sequences of 13 brown algae representing six phaeophycean orders: Dictyotales, Ectocarpales, Fucales, Laminariales, Sphacelariales, and Syringodermatales. These 15 brown algae included 10 ectocarpoids representing nine ectocarpacean (sensu Gabrielson et al. 1989) families: Chordariaceae, Dictyosiphonaceae, Ectocarpaceae, Elachistaceae, Heterochordariaceae, Leathesiaceae, Punctariaceae, Ralfsiaceae, and Scytosiphonaceae. We addressed the controversial taxonomic placement of A. japonicus and R. fungiformis in the Ectocarpales by analysis of DNA sequences. Neighbor-joining and maximum parsimony-inferred phylogenies provided evidence that A. japonicus and R. fungiformis are not closely associated with the other representatives of the Ectocarpales. Bootstrap analyses suggest polyphyly of the order Ectocarpales (sensu Gabrielson et al.). However, our analysis failed to resolve the phylogenetic relationship between A. japonicus and R. fungiformis. Our results suggest that the ectocarpoids are just as distantly related to A. japonicus and R. fungiformis as they are to members of the advanced orders Desmarestiales, Dictyotales, Fucales, Laminariales, Sphacelariales, and Syringodermatales.