“DNA signaturing” database construction for Tetradesmus species identification and phylogenetic relationships of Scenedesmus-like green microalgae (Scenedesmaceae,Chlorophyta)

Species identification of Scenedesmus-like microalgae, comprising Desmodesmus, Tetradesmus, and Scenedesmus, has been challenging due to their high morphological and genetic similarity. After developing a DNA signaturing tool for Desmodesmus identification, we built a DNA signaturing database for Tetradesmus. The DNA signaturing tool contained species-specific nucleotide sequences of Tetradesmus species or strain groups with high similarity in ITS2 sequences. To construct DNA signaturing, we collected data on ITS2 sequences, aligned the sequences, organized the data by ITS2 sequence homology, and determined signature sequences according to hemi-compensatory base changes (hCBC)/CBC data from previous studies. Four Tetradesmus species and 11 strain groups had DNA signatures. The signature sequence of the genus Tetradesmus, TTA GAG GCT TAA GCA AGG ACCC, recognized 86% (157/183) of the collected Tetradesmus strains. Phylogenetic analysis of Scenedesmus-like species revealed that the Tetradesmus species were monophyletic and closely related to each other based on branch lengths. Desmodesmus was suggested to split into two subgenera due to their genetic and morphological distinction. Scenedesmus must be analyzed along with other genera of the Scenedesmaceae family to determine their genetic relationships. Importantly, DNA signaturing was integrated into a database for identifying Scenedesmus-like species through BLAST.     

Physiological,Biochemical, and Molecular Characters for the Taxonomy of the Subgenera of Scenedesmus (Chlorococcales,Chlorophyta)

Biochemical and physiological properties of 16 Scenedesmus species representing the three subgenera Scenedesmus, Acutodesmus, and Desmodesmus are not suitable for species differentiation. All Scenedesmus species studied thus far produce secondary carotenoids, e.g. astaxanthin and canthaxanthin, under nitrogen-deficient conditions. In addition, with the exception of one strain, hydrogenase activity under anaerobic conditions is generally present. Sequence analyses of ribosomal 18S RNAs indicate that the subgenus Desmodesmus is phylogenetically well separated from the other subgenera, whereas the separation of Scenedesmus and Acutodesmus appears less convincing and is dismissed in favour of a single subgenus, Scenedesmus. Three taxa formerly assigned to the genus Chlorella are shown to be unicellular species of the genus Scenedesmus. “Chlorella” fusca var. vacuolata and “C.” fusca var. rubescens, which is closely related to S. obliquus, belong to the Scenedesmus/‘Acutodesmus group. “C.” fusca var. fusca is closely related to S. communis and thus belongs to the subgenus Desmodesmus. Inclusion of Kermatia pupukensis into the genus Scenedesmus is also strongly supported by the RNA data which furthermore indicate a relationship with the subgenus Desmodesmus.     

INDUCIBLE COLONY FORMATION WITHIN THE SCENEDESMACEAE: ADAPTIVE RESPONSES TO INFOCHEMICALS FROM TWO DIFFERENT HERBIVORE TAXA1

We studied the occurrence of colony formation within 40 different strains of Scenedesmaceae (Chlorococcales, Chlorophyta) in response to grazing‐released infochemicals from the herbivorous zooplankters Brachionus calyciflorus Pallas (Rotifera) and Daphnia magna Strauss (Cladocera). With the exception of two strains, all strains showed similar responses to both B. calyciflorus and D. magna infochemicals, either no response or inducible colony formation. Colony size was found to increase with B. calyciflorus infochemical concentration and could be described by a sigmoid function. The increase in colony size was more pronounced in the Scenedesmus species tested than in Desmodesmus species, which was probably due to higher threshold infochemical concentrations for colony induction in Desmodesmus. Therefore, the adaptivity of colony formation to the herbivory threat only holds above the threshold concentration for colony induction and as long as maximum colony size has not been attained. Taking this into account, our results suggest that inducible colony formation is a common adaptive response of many Scenedesmaceae to the threat of herbivory.     

Physiological and biochemical contributions to the taxonomy of the genera Ankistrodesmus and Scenedesmus

The genera Ankistrodesmus (including Raphidium) (17 strains) and Scenedesmus (27 strains) have a rather low salt tolerance without distinct speciesspecific differences. Only one strain of Ankistrodesmus and 23 strains of Scenedesmus are able to grow in the presence of 1% NaCl. Members of these genera therefore appear to be rather unsuitable for mass cultures in media based on sea water.-Two strains of Scenedesmus are thermophilic and grow well at 38°C.     

Physiological and biochemical contributions to the taxonomy of the genera Ankistrodesmus and Scenedesmus

The 17 Ankistrodesmus strains studied do not produce extracellular amylolytic enzymes. 20 out of 28 Scenedesmus strains, however, are able to hydrolyse starch.The groups of strains previously characterized physiologically and biochemically were assigned to established species of the genera Ankistrodesmus (including Monoraphidium) and Scenedesmus.Abbreviation GC Guanine + cytosine     

Phylogenetic inferences in Antennaria (Asteraceae: Gnaphalieae: Cassiniinae) based on sequences from Nuclear Ribosomal DNA internal transcribed spacers (ITS)

The phylogenetic relationships among sexually reproducing species of Antennaria (Asteraceae) are poorly understood. An earlier cladistic analysis based on morphology did not fully resolve the phylogeny of these taxa and therefore a different approach using molecular data was explored. The internal transcribed spacer regions (ITS-1 and ITS-2) of nuclear ribosomal DNA were sequenced for 30 species of Antennaria and one species from each of the outgroup genera Anaphalis, Ewartia, Leontopodium, and Pseudognaphalium. The ITS-1 sequence in Antennaria ranged from 253 to 260 base pairs (bp) in length, and the proportion of nucleotide differences between pairs of species of Antennaria ranged from 1 to 14%. For ITS-2, the divergence between pairs of species of Antennaria ranged from 0 to 8%. ITS-2 is shorter than ITS-1, ranging from 213 to 219 bp. Phylogenetic analysis indicates that, relative to the outgroups included, Antennaria is a well-supported monophyletic group. Based on the genera surveyed, Leontopodium appears to be the sister genus of Antennaria. The general topology of the molecular trees agrees with that based on previous morphological analyses and indicates that Antennaria is composed of six clades of equal rank, corresponding to the traditionally recognized informal groups, the Geyeriae, Argenteae, Arcuatae, Dimorphae, Pulcherrimae, and Catipes. Sequence and morphological data indicate that the Alpinae and Dioicae are unnatural, polyphyletic units that should be abandoned and redefined as the monophyletic Catipes group. Phylogenetic analysis of ITS sequences also suggests the dissociation of A. stenophylla from the Dimorphae, where it is traditionally placed, and its affiliation with the Argenteae, as well as the placement of A. arcuata in its own group.     

Cell wall composition of chlorococcal algae

The cell walls of representatives of the genera Chlorella, Monoraphidium, Ankistrodesmus and Scenedesmus contained 24–74% neutral sugars, 1–24% uronic acids, 2–16% protein and 0–15% glucosamine. Two types of cell walls could be discerned containing as main sugars either rhamnose and galactose or mannose and glucose with a lack of galactose.     

Sequence variation in ITS spacers and 5.8S rDNA and relationship of E, St, P, Ns, Xm, and H genomes in the genera of Agropyron, Elytrigia, Leymus, Pascopyrum, Psathyrostachys, and Hordeum

Understanding species evolution and improvement requires information of their genome origin and differentiation. Among the species in the family Gramineae, genome identities of Agropyron-Elytrigia-Leymus group are still ambiguous. In order to delineate the genome relationship, nucleotide sequence analysis in the rDNA ITS regions was carried out among the species in the genera Elytrigia, Agropyron, Psathyrostachys, Leymus, and Psacopyrum containing E, St, P, Ns, and Xm genomes. The ITS-1 and ITS-2 showed a narrow range of variation in length except for the presence of a pentanucleotide, TGGGG, in/del in some haplotypes, whereas higher numbers of nucleotide substitutions were observed in most genera. There were 187 variable sites in the ITS-1, 5.8S, and ITS-2 regions, in which a few genome specific mutations were observed. While the level of variation was similar between ITS-1 and ITS-2, the rate of transition mutation versus transversion mutations was different among the ITS-1, 5.8S, and ITS-2 segments. GC contents of the ITS regions ranged between 55–65% between genomes and the haplotypes of P and H genomes were slightly higher than others. In phylogenetic analysis, the ITS haplotypes were classified into two groups; one containing H, Ns, NsXm genomes, and another containing P, St, and E genomes, which are congruous to the genome affinities from other studies. Among the four genomes in Pascopyrum smithii (2n=8x=56, StStNsNsHHXmXm), the haplotypes of H and St genomes were identified with the reference diploid species, but the haplotypes having Ns and Xm genomes were not found in the present analysis.     

The mechanism of hydrogen photoproduction by several algae

Summary The contribution of PS II to H₂ photoproduction by several unicellular green algae was measured both when O₂ evolution and photophosphorylation were unimpaired and also when these processes had been eliminated by Cl-CCP. As judged by the effects of DCMU, a PS II contribution was found under both sets of experimental conditions for several strains of Chlorella, Ankistrodesmus and Scenedesmus. However, H₂ photoproduction by Chlamydomonas moewusii was insensitive to DCMU and thus was entirely due to PS I. With cells treated with Cl-CCP, the relative amount of PS II contribution varied from zero in autotrophically grown Chlamydomonas reinhardii, to 20% in photoheterotrophically grown and 50% in autotrophically grown cells of Ankistrodesmus braunii, Chlorella fusca, Chlorella vulgaris and Scenedesmus obliquus. The dehydrogenation of reduced H-donors by PS II of Scenedesmus treated with Cl-CCP showed the same biphasic kinetics previously described for H₂ photoproduction by PS I of this alga.Abbreviations Cl-CCP carbonyl cyanide m-chlorophenylhydrazone - DCMU 3-(3,4-dichlorophenyl)-1,1-dimethylurea - ICC Indiana Culture Collection - PS photosystem - SAL salicylaldoxime - SIO Marine Botany Culture Collection, Scripps Institution of Oceanography These studies were supported by contract No. AT-(40-1)-2687 from the U.S. Atomic Energy Commission.     

Phylogenetic relationships of Nemania plumbea sp. nov. and related taxa based on ribosomal ITS and RPB2 sequences

During a survey fungal diversity of xylariaceous fungi in Thailand, a new Nemania species, N. plumbea, was identified. Nemania plumbea is characterized by soft-textured grey stromata on a persistent mat of white hyphae, pale brown ascospores with a short germ-slit on the more convex side. It also produces a Geniculosporium-like anamorph in culture. In order to evaluate its phylogenetic relationships among related species and genera, ITS-5.8S rDNA and RPB2 were analysed separately and simultaneously. Results from the phylogenetic analyses indicate that there is close phylogenetic association between N. plumbea and N. aenea. A preliminary account into the natural grouping of Xylariaceae based on ITS-5.8S rDNA and RPB2 sequences is also discussed.     

Hydrogen evolution by several algae

Summary Out of 33 strains of unicellular algae examined, H₂ evolution was observed only in species of Chlamydomonas, Chlorella and Scenedesmus. While the photoevolution of H₂ by these algae was generally stimulated both by an organic substrate and by the uncoupler CCCP¹, response to DCMU varied. On the basis of the response to DCMU, it was concluded that the mechanism of photoevolution of H₂ differed from one alga to another. The reaction in some algae appeared to be dependent on either the photooxidation of water or oxidative carbon metabolism for reductant; that in other algae was supported by reductant from both these sources.     

Ribosomal DNA ITS-1 Intergenic Spacer Polymorphism in Triatomines (Triatominae, Heteroptera)

The length polymorphism of ribosomal DNA ITS-1 intergenic spacer was analyzed in eight species of triatomines belonging to Triatoma, Rhodnius, and Panstrongylus genera. The analyzed species were Rhodnius domesticus, R. neivai, R. robustus, Triatoma brasiliensis, T. infestans, T. vitticeps, Panstrongylus megistus, and P. herreri. These insects are vectors of Chagas' disease, one of the most prominent public health problems among South American countries. This work allowed the differentiation between species of the Triatomini and Rhodniini tribes through the analysis of ITS-1 length polymorphism by PCR and RFLP techniques. The species of the Triatoma and Panstrongylus genera presented an amplified ITS-1 fragment between 600 and 1000 bp, whereas Rhodnius presented a less variable ITS-1 length fragment, around 300 bp, which could reflect the monophyletic origin of the Rhodniini tribe. Species belonging to this genus were further differentiated by RFLP with HaeIII and AluI endonucleases. Our results corroborate the hypothesis of polyphyletic origin in this group of insects and contribute to knowledge about evolutionary relationships in triatomines.     

Molecular phylogenetic studies of Lachnum and its allies based on the Japanese material

Molecular phylogenetic studies were carried out based on ITS-5.8S rDNA, the D1–D2 region of the large subunit rRNA gene, RPB2, and combined data of D1–D2 and RPB2 as well as these three genes on 36 species among 7 genera for Lachnum and allied genera in the family Hyaloscyphaceae. In the combined data of all three regions, seven strongly supported clades were obtained. The same clades were also recognized in most of the trees based on each gene, and the combined data of D1–D2 and RPB2, although some of them were not strongly supported. Four clades represented Albotricha, Brunnipila, Incrucipulum, and Lachnellula, respectively, whereas Lachnum was distributed to the remaining three clades. The molecular phylogenies strongly supported a group of species with granulate hairs, and we suggest the concept of Lachnaceae should be restricted to these species. Based on the molecular phylogenetic analysis, three new combinations—Incrucipulum longispineum, I. radiatum, and Lachnellula pulverulentum from Lachnum—are proposed.     

Über das ätherische Öl von Grünalgen

Zusammenfassung Das ätherische Öl und seine Hauptkomponenten (Proazulene) erwiesen sich auch bei den Gattungen Ankistrodesmus und Scenedesmus als brauchbare taxonomische Merkmale. Der Ölgehalt licgt in der gleichen Größenordnung wie innerhalb der Gattung Chlorella. Einige Ankistrodesmus-Stämme, die keine Proazulene bilden, müssen offensichtlich anderen Gattungen zugeordnet werden.

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On the essential oil of green algaeII. The oils of the genera Ankistrodesmus and Scenedesmus

The essential oil and its main components (i.e., proazulenes) are useful taxonomic characters also in the genera Ankistrodesmus and Scenedesmus. The amounts of oil in these genera are similar to that of the genus Chlorella. A few strains of Ankistrodesmus, which are unable to synthesize proazulenes, seem to belong to other genera.

     

DNA base composition within the genusScenedesmus (Chlorophyta)

The DNA base compositions of 60 strains ofScenedesmus were determined and found to be a valuable indicator for the differentiation within this genus (except for the very closely related species of the subsect.Desmodesmus). The range for allScenedesmus species was 49.9–69.3 mol% GC for nuclear DNA and 36.8–39.9 mol% GC for chloroplast DNA. The separation of the genusTetradesmus cannot be verified by GC values, becauseS. obliquus andS. (Tetradesmus)wisconsinensis have a similar GC content.S. (Chlorella)ultrasquamata, S. costato-granulatus (sect.Costato-granulati) andS. lunatus are separated clearly from all other species of the genus because of their high GC content.     

Importance of Nutrient Competition and Allelopathic Effects in Suppression of the Green Alga Scenedesmus obliquus by the Macrophytes Chara, Elodea and Myriophyllum

Possible allelopathic effects of substances released from the macrophytes Chara globularis, Elodea canadensis, Myriophyllum spicatum on the common green alga Scenedesmus obliquus were tested in the laboratory with plastic plants and untreated medium as controls. A two-phase approach was used in which first the effects of physical presence of plants was studied (phase I) followed by the effects of plant culture filtrates (phase II). In the presence of plastic plants growth was reduced only marginally, but strong growth inhibition of Scenedesmus occurred in the physical presence of all macrophytes. In contrast, filtrates from Chara had no growth inhibitory effect on Scenedesmus. Myriophyllum filtrate reduced particle-based growth rate by 7% compared to filtration controls, while Elodea culture filtrate reduced volume-based growth by 12%, chlorophyll-based growth by 28% and particle-based growth by 15%. Photosystem II-efficiency of Scenedesmus was reduced in all three macrophyte treatments in phase I, but not in filtrates from macrophyte cultures (phase II). Thus, while enzyme activity or other physiological aspects may have been affected, the current study yielded no proof for allelopathically active compounds being directed at photosynthesis. Mean particle volume (MPV) of Scenedesmus was not influenced by macrophyte exudates and cultures remained dominated by unicells. The strong growth inhibitory effects found for Scenedesmus in the physical presence of macrophytes, but not in plastic controls, and no or weaker response in nutrient-enriched filtrates, suggest nutrient competition was a more powerful driving factor than allelochemicals. However, the experimental design does not exclude disappearance of allelochemicals during the filtration process.     

Molecular phylogeny of a green microalga isolated from White Sea sponge <Emphasis Type="Italic">Halichondria panicea</Emphasis> (Pallas, 1766)

Molecular identification of eukaryotic microalga 1Hp86E-2 isolated from White Sea sponge Halichondria panicea (Pallas, 1766) was conducted, and phylogenetic analysis was carried out using the nucleotide sequence of 18S rRNA gene (GenBank, no. JX437624). Isolated microalga was classified to the genus Desmodesmus. Microalga 1Hp86E-2 proved to be closely related to the algae Desmodesmus sp. 3Dp86E-1, Desmodesmus sp. 2C166E, and Desmodesmus sp. 1Pm66B isolated from White Sea invertebrates. Phylogenetic analysis showed that these closely related organisms belong to a monophyletic group.     

Observations over 24 hours on the quantity of algae inside grazing ciliated protozoa

Summary The numbers of cells of the alga Scenedesmus (S. quadricauda and S. denticulatus) inside natural populations of grazing ciliated protozoa from a eutrophic pond (Loxodes magnus and L. striatus) were counted at 4 h intervals over two 24 h periods. There were no diel fluctuations in number of algal cells inside the protozoa, therefore the grazing rates were probably more or less constant over 24 h. L. magnus contained more algae than the smaller L. striatus. S. quadricauda was the more abundant Scenedesmus species in the phytoplankton and both Loxodes species contained more S. quadricauda than S. denticulatus, although the ratio S. denticulatus cells/S. quadricauda cells was greater inside the ciliates than in the phytoplankton.     

Hydrocarbons in green and blue-green algae

Liquid column chromatography and thin-layer chromatography were used to determine the total content of hydrocarbons and gas chromatography was used to evaluate composition of hydrocarbons in green algae (Chlorella kessleri, C. vulgaris, Chlorella sp.,Scenedesmus acutus, S. acuminatus, S. obliquus) and the blue-green alga (Spirulina platensis) cultivated under autotrophic or heterotrophic conditions. InC. kessleri cultivated under heterotrophic conditions the content of hydrocarbons was found to be about 10^-2 % (per dry mass), whereas under autotrophic conditions it was about 10^-3 % (per dry mass). The highest content of hydrocarbons was detected in species of the genusScenedesmus cultivated autotrophically (10^-1 %). Heptadecane and hexacosane were found as major alkanes, 1-heptadecene was detected among alkenes.