Structure of the Large Subunit rDNA from a Diatom, and Comparison Between Small and Large Subunit Ribosomal RNA for Studying Stramenopile Evolution

The aim of this study was to compare the usefulness of complete small and large subunit rRNA, and a combination of both molecules, for reconstructing stramenopile evolution. To this end, phylogenies from species of which both sequences are known Acre constructed with the neighbor-joining, maximum parsimony, and maximum likelihood methods. Also the use of structural features of the rRNAs was evaluated. The large subunit rRNA from the diatom Skeletonema pseudocostatum was sequenced in order to have a more complete taxon sampling, and a group I intron was identified. Our results indicated that heterokont algae are monophyletic, with diatoms diverging first. However, as the analysis was restricted to a particular data set containing merely six taxa, the outcome has limited value for elucidating stramenopile relationships. On the other hand, this approach permits comparison of the performance of both rRNA molecules without interference from other factors, such as a different species selection for each molecule. For the taxa used, the large subunit rRNA clearly contained more phylogenetic information than the small subunit rRNA. Although this result can definitely not be generalized and depends on the phvlogeny to be studied, in some cases determining complete large subunit rRNA sequences certainly seems worthwhile.     

Phylogeny of the Bangiophycidae (Rhodophyta) and the secondary endosymbiotic origin of algal plastids

The Rhodophyta (red algae) are composed of the subclasses Bangiophycidae and Florideophycidae. Two evolutionarily interesting features of the Bangiophycidae are: (1) they are the ancestral pool from which the more morphologically complex taxa in the Florideophycidae have arisen and (2) they are the sources of the plastids, through secondary endosymbioses, for the Cryptophyta, Haptophyta, and the Heterokonta. To understand Bangiophycidae phylogeny and to gain further insights into red algal secondary endosymbioses, we sequenced the plastid-encoded small subunit ribosomal DNA (rDNA) coding region from nine members of this subclass and from two members of the Florideophycidae. These sequences were included in phylogenetic analyses with all available red algal plus chlorophyll a + c algal plastid rDNA coding regions. Our results are consistent with a monophyletic origin of the Florideophycidae with these taxa forming a sister group of the Bangiales. The Bangiophycidae is of a paraphyletic origin with orders such as the Porphyridiales polyphyletic and distributed over three independent red algal lineages. The plastids of the heterokonts are most closely related to members of the Cyanidium-Galdieria group of Porphyridiales and are not directly related to cryptophyte and haptophyte plastids. The phylogenies provide strong evidence for the independent origins of these "complex" algal plastids from different members of the Bangiophycidae.     

The evolution of the diatoms (Bacillariophyta). I. Origin of the group and assessment of the monophyly of its major divisions

The diatoms are one of the best characterised algal groups. Despite this, little is known of the evolution of the group from the earliest cell to the myriad of taxa known today. Relationships among taxa at the family or generic level have been recognised in some diatoms. However, relationships at higher taxonomic levels are poorly understood and have often been strongly influenced by the first appearances of key taxa in the fossil record. An independent assessment of relationships among the diatoms at these higher taxonomic levels has been made using rRNA sequence data to infer phylogenetic relationships. In this paper we present an analysis of 18S rRNA data from several chosen centric, araphid and raphid pennate taxa. The phylogenetic inferences from these 18S rRNA sequences are supported by evidence from the fossil record and evidence from ontogenetic data. Ribosomal RNA data indicate that both the centric and araphid pennate lineages may not be monophyletic.     

Diversity In Symbiotic Dinoflagellates (Pyrrhophyta) from Seven Scleractinian Coral Species: Restriction Enzyme Analysis of Small Subunit Ribosomal RNA Genes

ABSTRACT The diversity of symbiotic dinoflagellates (SD) from seven coral species ( Fungia scutaria, Fungia paumotensis, Lep-tastrea transversa, Pavona cactus, Pocillopora verrucosa, Montastrea curia , and Acropora fonnosa ) was studied in a restricted geographical area, the Lagoon of Arue on the island of Tahiti. Their diversity was explored by small subunit ribosomal RNA gene (SSU rDNA) restriction fragment length polymorphism (RFLP). After a nested amplification with SD specific primers, RFLP analyses were performed directly and after a cloning step. The diversity of these different SSU rDNA was estimated in respect to possible technical artifacts. In an axenic culture of SD from the coral Galaxea fascicularis , both heterogeneous SSU rDNAs and artifact molecules were observed as in our SD samples. According to the number of patterns observed, corals Fungia paumotensis, Leptastrea transversa. Pavona cactus, Montastrea curia, and Acropora fonnosa contained one class of SD SSU rDNAs. whereas Fungia scutaria and Pocillopora verrucosa contained three and two classes of SD SSU rDNAs respectively. In the limited geographic area studied. SD from different coral species shared the same pattern, except SD from Montastrea curta , which showed a unique pattern. In addition to the possibility of SD flux among different coral species, specific mechanisms could also be involved in the establishment of a symbiosis.     

Functional Study of the Residue C899 in the 900 Tetraloop of Escherichia coli Small Subunit Ribosomal RNA

A mutant ribosome bearing C899G in the 900 tetraloop of Escherichia coli 16S rRNA, one implicated in a conformational switch in the dynamic movements of the ribosome, showed defects in subunit association and 30S initiation complex formation. Our results explain the basis of the loss of protein synthesis ability caused by a perturbation of the 900 tetraloop.     

The Evolution of the Vahlkampfiidae as Deduced from 16S-like Ribosomal RNA Analysis

ABSTRACT. The amoebae, a phenotypically diverse, paraphyletic group of protists, have been largely neglected by molecular phy-logeneticists. To better understand the evolution of amoebae, we sequenced and analyzed the 16S-like ribosomal RNA genes of three vahlkampfiid amoebae: Paratetramitus jugosus, Tetramitus rostratus and Vahlkampfia lobospinosa . The Vahlkampfiidae lineage is monophyletic, branches early along the eukaryotic line of descent, and is not a close relative of the multicellular amoebae that also reversibly transform from amoebae to flagellates.     

Serial replacement of a diatom endosymbiont in the marine dinoflagellate Peridinium quinquecorne (Peridiniales, Dinophyceae)

To infer the phylogeny of both the host and the endosymbiont of Peridinium quinquecorne Abé, the small subunit (SSU) ribosomal DNA (rDNA) from the host and two genes of endosymbiont origin (plastid‐encoded rbcL and nuclear‐encoded SSU rDNA) were determined. The phylogenetic analysis of the host revealed that the marine dinoflagellate P. quinquecorne formed a clade with other diatom‐harbouring dinoflagellates, including Kryptoperidinium foliaceum (Stein) Lindeman, Durinskia baltica (Levander) Carty et Cox and Galeidinium rugatum Tamura et Horiguchi, indicating a single endosymbiotic event for this lineage. Phylogenetic analyses of the endosymbiont in these organisms revealed that the endosymbiont of P. quinquecorne formed a clade with a centric diatom (SSU data indicated it to be closely related to Chaetoceros), whereas the endosymbionts of other three dinoflagellates formed a clade with a pennate diatom. The discrepancy between the host and the endosymbiont phylogenies suggests a secondary replacement of the endosymbiont from a pennate to a centric diatom in P. quinquecorne.     

A molecular and ultrastructural description of Spathidiopsis buddenbrocki and the phylogenetic position of the family Placidae (Ciliophora)

Spathidiopsis and Placus are the only two genera within the family Placidae. The family has been placed in the class Prostomatea and order Prorodontida because its members have somatic monokinetids with a radial transverse ribbon, a straight non‐overlapping postciliary ribbon, and anteriorly directed non‐overlapping kinetodesmal fibril, an apical cytostome lacking specialized oral cilia, a brosse, and toxicysts. To confirm the stability of this placement, ultrastructural morphology and small subunit rRNA gene sequences of Spathidiopsis socialis, Spathidiopsis buddenbrocki, and Placus striatus were determined. These data were combined with information from other ciliates, and phylogenetic trees were generated using maximum‐likelihood and maximum‐parsimony methods. The analyses confirmed the family Placidae to be a monophyletic group in the Prostomatea with the Placidae a sister group to a Cryptocaryon + Coleps + Prorodon clade.     

Phylogeny of the Poorly Known Ciliates,Microthoracida, a Systematically Confused Taxon (Ciliophora), with Morphological Reports of Three Species

Three species of Microthoracids, Lopezoterenia paratorpens n. sp., Trochiliopsis australis Foissner et al., 1988 and Discotricha papillifera Tuffrau, 1954, collected from Chinese coastal waters, were investigated using live observation and protargol staining methods. Lopezoterenia paratorpens n. sp. was characterized by its squarely shaped cortical papillae and by dorsal kineties which contained loosely distributed basal bodies. Trochiliopsis australis was revealed to have two oral membranelles, which was not recorded in the original report. Phylogenetic analyses were carried out based on SSU rRNA gene sequence data from each of the three species, and on other available data for microthoracids. The results showed that the order Microthoracida is not monophyletic because the family Discotrichidae, which contains L. paratorpens and D. papillifera, forms a clade separated from the “core” Microthoracids clade. The topologies of the maximum likelihood and Bayesian inference trees, along with the distinct morphological characteristics found previously, suggest that the family Discotrichidae should not be assigned to the order Microthoracida. We propose to designate a new order, Discotrichida n. ord. which diagnosed as: flattened ciliates with conspicuous cortical papillae on both dorsal and ventral faces, rod‐shaped mucocysts, and an asymmetric cytopharyngeal basket. Also, the fact that Leptopharynx clusters with the assemblage including T. australis, and Pseudomicrothorax is located distantly from Leptopharynx indicates that the classification of Pseudomicrothoracidae and Microthoracidae by Foissner (1985) is justified.     

Description of Paratetrahymena parawassi n. sp. using Morphological and Molecular Evidence and a Phylogenetic Analysis of Paratetrahymena and Other Taxonomically Ambiguous Genera in the Order Loxocephalida (Ciliophora,Oligohymenophorea)

ABSTRACT. The marine scuticociliate Paratetrahymena parawassi n. sp. is described on the basis of morphology, especially infraciliature, and the sequence of its small subunit (SSU) rRNA gene to become the second known member of its genus. Paratetrahymena and other ciliates in the order Loxocephalida possess a mixture of morphological and morphogenetic features characteristic of the subclasses Hymenostomatia and Scuticociliatia. Accordingly, we used SSU rRNA sequences to analyze the phylogeny of Paratetrahymena and three other loxocephalid genera. Paratetrahymena and Cardiostomatella vermiformis formed a moderately well‐supported clade that diverged at a deep level from all other scuticociliates, supporting separation of loxocephalids from other scuticociliates as a suprafamilial taxon. Sathrophilus holtae was a sister taxon to Paratetrahymena and Cardiostomatella in a poorly supported, unresolved relationship; nevertheless, association of all three genera into a single clade was supported by an approximately unbiased (AU) test. Any association of these genera singly or as a group with the Hymenostomatia was rejected decisively by AU tests and by a complete absence in the loxocephalids of the unique nucleotide identities that distinguish hymenostomes. Therefore, the morphological and morphogenetic similarities of loxocephalids to hymenostomes may be plesiomorphies, and the conflicting mix of scuticociliate and hymenostome characteristics seen in loxocephalids may result from differing rates of character evolution. Dexiotrichides pangi and Urocentrum, which is currently classified as a peniculid, formed a small clade that associated with hymenostomes and peritrichs. Monophyly of the Loxocephalida with Dexiotrichides and/or Urocentrum included was not rejected by AU; however, inclusion of Urocentrum in the Peniculia was rejected by AU tests. A hypothesis is offered to explain the lack of resolution of loxocephalid ciliates and Urocentrum in phylogenetic trees, namely that their phylogenetic positions are influenced by a combination of heterogeneous data and long‐branch attraction caused by poor representation of taxa in analyses. The well‐known genus Cyclidium, a member of the order Pleuronematida, was revealed to be polyphyletic as a byproduct of our analyses of loxocephalids. In particular, Cyclidium porcatum appears to fall outside the clade containing typical members of the subclass Scuticociliatia and thus invites investigation as a possible member of the order Loxocephalida.     

A Conserved Motif in the 5.8S Ribosomal RNA (rRNA) Gene is a Useful Diagnostic Marker for Plant Internal Transcribed Spacer (ITS) Sequences

The nuclear ribosomal DNA (rDNA) internal transcribed spacer (ITS) region has become an important nuclear locus for molecular systematic investigations of angiosperms at the intergenic and interspecific levels. Universal PCR primers are positioned on the conserved rRNA genes (18S, 5.8S, 26S) to amplify the entire ITS spacer region. Recent reports of fungal and algal contaminants, first described as plant ITS sequences, stress the need for diagnostic markers specific for the angiosperm ITS region. This report describes a conserved 14 base pair (bp) motif in the 5.8S rRNA gene that can be used to differentiate between flowering plants, bryophytes, and several orders of algae and fungi, including common plant pathogenic and non-pathogenic fungi. A variant of the motif (found in fungi and algae) contains a convenient EcoRI restriction site that has several applications for eliminating problematic contaminants from plant ITS preparations.     

Expanded Phylogenetic Representation of Genera Opercularia and Epistylis Sheds Light on the Evolution and Higher‐Level Taxonomy of Peritrich Ciliates (Ciliophora: Peritrichia)

ABSTRACT. We have generated 18S rRNA sequences for peritrichs collected in Brazil, including four Opercularia species, two different populations of Epistylis plicatilis (one epibiont and another free‐living), and one additional Epistylis species. Our Opercularia species clustered with the previously available Opercularia microdiscum, corroborating the monophyly of this genus. The Epistylis sampled here clustered with previously sequenced species of this genus. The two populations of E. plicatilis collected in Brazil clustered closely together despite their different ecological contexts, whereas both were very divergent from the sample assigned to the same species previously sampled in China. If affirmed by additional morphological corroboration of species assignment, this observation would indicate that samples from different continents morphologically allocated in the same species may in fact belong to distant evolutionary lineages. More broadly, our results support the recognition of two major clades within Peritrichia. Given the robustness of their support, we suggest that these two clades should be formally recognized as orders, and propose the names Vorticellida and Operculariida to designate them. Furthermore, Epistylis species occurred in both orders, tending to occupy basal positions. This suggests that characters used to define this genus may be plesiomorphic for Peritrichia, so that Epistylis may in fact represent an assemblage of basal species retaining ancestral features.     

Morphologic and molecular data suggest that Lynnella semiglobulosa n. g., n. sp. represents a new family within the subclass Choreotrichia (Ciliophora, Spirotrichea)

The morphology, morphogenesis, and phylogeny of an undescribed oligotrich sensu lato (s. lat.) ciliate, Lynnella semiglobulosa n. g., n. sp., found in Daya Bay, southern China, were investigated. This species shares some features with both oligotrichs sensu stricto and choreotrichs, but most morphological and morphogenetic characters as well as the phylogenetic analysis suggest that it should be assigned into subclass Choreotrichia temporarily. Lynnella semiglobulosa is distinguished from members of all known genera and families of the subclass Choreotrichia by a unique combination of characteristics of the buccal and somatic ciliatures. Thus, a new family Lynnellidae n. fam. and a new genus Lynnella n. g. are proposed for it. The new family is distinguished by an open adoral zone of membranelles (AZM) in which, however, there are no ventral membranelles; the distal and proximal portions of the new adoral zone lie close to each other forming an open circle in stomatogenesis. The new genus Lynnella is characterized by possessing two longitudinally oriented somatic kineties, one dorsal and one ventral, several proximal membranelles progressively lengthened toward the proximal end of adoral zone, and two macronuclear nodules. In phylogenetic analyses based on small subunit rRNA gene sequences, L. semiglobulosa clustered basally to all choreotrichs, but with relatively weak support; nevertheless, the possibility of a relationship with the subclass Oligotrichia was not rejected by the approximately unbiased nor Shimodaira-Hasegawa test. Based on morphological, morphogenetic, and molecular evidence of L. semiglobulosa, it is confirmed that the open AZM should be a plesiomorphic character of oligotrichs s. lat. as suggested previously.     

Evolutionary relationships among heterokont algae (the autotrophic stramenopiles) based on combined analyses of small and large subunit ribosomal RNA

In order to study the phylogenetic relationships within the stramenopiles, and particularly among the heterokont algae, we have determined complete or nearly complete large-subunit ribosomal RNA sequences for different species of raphidophytes, phaeophytes, xanthophytes, chrysophytes, synurophytes and pinguiophytes. With the small- and large-subunit ribosomal RNA sequences of representatives for nearly all known groups of heterokont algae, phylogenetic trees were constructed from a concatenated alignment of both ribosomal RNAs, including more than 5,000 positions. By using different tree construction methods, inferred phylogenies showed phaeophytes and xanthophytes as sister taxa, as well as the pelagophytes and dictyochophytes, and the chrysophytes/synurophytes and eustigmatophytes. All these relationships are highly supported by bootstrap analysis. However, apart from these sister group relationships, very few other internodes are well resolved and most groups of heterokont algae seem to have diverged within a relatively short time frame.     

PCR-Based Ribosomal DNA Detection Technique for Microalga (Heterosigma carterae) Causing Red Tide and Its Application to a Biosensor Using Labeled Probe

A technique for detecting Raphidophycean, a bloom-forming genus of algae, was developed using a specific DNA probe. The design of the probe was based on a sequence polymorphism within the small subunit (SSU) ribosomal RNA gene (rDNA) of this strain by using fluorescence polarization (FP) analysis and the BIAcore 2000 biosensor, which utilized surface plasmon resonance (SPR). The specific sequence in SSU rDNA for Heterosigma carterae was determined by sequence data analysis. One pair of polymerase chain reaction (PCR) probes was designed for use in making the identification. H. carterae SSU rDNA was amplified by PCR. Using a fluoroscein isothiocyanate–labeled or biotin-labeled oligonucleotide probe, the PCR-amplified rDNA was selectively detected as an FP-intensity change via FP analysis or as a resonance-unit change via SPR. Although total time for final detection after sampling was within 3 hours, specific rDNA could be detected within 10 minutes after PCR through these detection methods.     

基于线粒体12S和16S rRNA基因部分序列探讨蚱科12种的系统发育关系

用ABI377自动测序仪测定了蚱科5属11个种的12s和16S rRNA基因部分序列,并从GenBank获得1属1种的同源序列;用Clustal X1．81比较其同源性,用Mega2．1计算序列变异性和遗传距离。在获得的736bp序列中,A T含量为71．2％～77．5％,平均为73．9％;G C含量为22．5％～28．8％,平均为26．1％。经Clustal X1．81软件比对,共得到755个位点,其中简约信息位点185个。以Cylindraustralia kochii为外群,构建NJ、MP和ML分子系统树,结果表明：(1)蚱属并非一个单系群,而是一个并系群;(2)环江柯蚱Coptltettix huanjiangensis和贡山柯蚱C.gongshanensis为同一个种,即贡山柯蚱,而环江柯蚱是贡山柯蚱的同物异名。