DISTINGUISHING MULTIPLE LINEAGES OF PEDIASTRUM DUPLEX WITH MORPHOMETRICS AND A PROPOSAL FOR LACUNASTRUM GEN. NOV1

Accurately defining species boundaries in the green algae (Chlorophyta) is integral for studies of biodiversity and conservation, water‐quality assessments, and the use of particular species as paleoindicators. Recent molecular phylogenetic and SEM analyses of the family Hydrodictyaceae (Chlorophyta) resolved three phylogenetic lineages of isolates with the Pediastrum duplex Meyen 1829 phenotype. The present study employed analyses of cell shape and cell wall ultrastructure to determine if the three lineages possessing the P. duplex morphotype were distinguishable. Only one of the groups, containing isolates with the P. duplex var. gracillimum West et G. S. West phenotype, was shown to be morphologically distinct from the other two P. duplex groups. The erection of a new genus, Lacunastrum, is proposed to recognize this group as a separate taxon.     

Growth and mineral nutrition of two Chlorococcalean algae

The effect of composition of the medium and pH on the growth of Pediastrum duplex Meyen and Dictyosphaerium pulchellum Wood was studied. Both species showed preference to alkaline conditions. The ammonium nitrate grown colonies of D. pulchellum lacked mucilage and showed a more compact form, resembling D. pulchellum var. minitum Deflandre. From this it appears that D. pulchellum var. minitum is a nutritional variant of the species and not a stable variety.     

MOLECULAR PHYLOGENETIC RELATIONSHIPS IN THE FRESHWATER FAMILY HYDRODICTYACEAE (SPHAEROPLEALES,CHLOROPHYCEAE), WITH AN EMPHASIS ON PEDIASTRUM DUPLEX1

The freshwater green algal family Hydrodictyaceae (Sphaeropleales, Chlorophyta) has traditionally consisted of four coenobial genera, Pediastrum Meyen 1829, Hydrodictyon Roth 1797, Sorastrum Kützing 1845, and Euastropsis Lagerheim1894. Two recent molecular phylogenetic studies demonstrated the need for reevaluation of the generic and species boundaries in this morphology‐rich family. This study expands the previous work to include phylogenetic analyses of 103 ingroup isolates representing North America, Europe, and Australia, with an emphasis on the common and geographically widespread species Pediastrum duplex. Nucleotide sequence data were collected from the nuclear LSU (26S rDNA) and the chloroplast RUBISCO LSU (rbcL) genes, totaling >3,000 aligned characters. The 26S and rbcL data sets were analyzed using maximum‐likelihood (ML) and Bayesian phylogenetic methods. In addition, SEM was used to examine the wall morphology of a majority of the isolates. The results supported previous indications that the P. duplex Meyen 1829 morphotype is nonmonophyletic and resolved some previously ambiguous relationships recovered in earlier phylogenetic estimations using fewer isolates. These new data allowed testing of the recent taxonomic revisions of the family that split Pediastrum into five genera. Some of the previous revisions by Buchheim et al. (2005) were well supported (erection of Stauridium and Monactinus), while others were not (Pediastrum, Pseudopediastrum, Parapediastrum).     

<Emphasis Type="Italic">Siderocelis irregularis</Emphasis> (Chlorophyta,Trebouxiophyceae) in Lake Tanganyika (Africa)

Siderocelis irregularis Hindák, representing a genus Siderocelis (Naumann) Fott that is known from European temperate waters, was identified as a common phytoplankter in Lake Tanganyika. It was found aposymbiotic as well as ingested (possibly endosymbiotic) in lake heterotrophs, mainly Strombidium sp. and Vorticella spp. The morphology and ultrastructure of the species, studied with LM, SEM and TEM, are described with emphasis on the structure of the cell wall and the pyrenoid. Presented at the International Symposium Biology and Taxonomy of Green Algae V, Smolenice, June 26–29, 2007, Slovakia.     

Growth characteristics of selected thermophilic strains of cyanobacteria using crossed gradients of temperature and light

Growth requirements of 10 possibly thermophilic strains of cyanobacteria were compared under a wide spectrum of light and temperature conditions (7–80Wm^?2, 12–40°C). The strains were isolated from different localities: six of thermal springs in Slovakia (4 from Pie??any, 2 from Sklené Teplice), one from thermal waters in Rupite, Bulgaria, 2 strains from a hypersaline lake Chott-el-Djerid, Tunisia, and one strain from the tropical island of Cebu, Philippines. Although the crossed gradient unit allowed only sub-optimal temperature range with respect to thermophile definition, i.e. optimum temperature above 45°C, there were difference among the strains. The most thermophilic and high-light tolerant strain was Synechococcus bigranulatus strain Lukavsky 2005/66, with a peak above 45°C; the second was Chroococcidiopsis thermalis strain Hindák 2008/9, and Isocystis sp. strain Hindák 2006/1. The temperature requirements of other strains were similar; the least thermophilic were both Slovakian strains of the genus Hapalosiphon. Growth was not limited below 80 W m^?2, except for H. fontinalis strain Hindák 2008/3, which prefers lower irradiance. Hapalosiphon delicatulus strain Hindák 2007/20, isolated from a bark tree on the tropical island of Cebu was rather mesophilic than thermophilic and shade-preferring. The CCA revealed that the ecologically similar strains originated from the same or nearby localities. There were no significant correlations between temperature optima in culture and in nature. Bulgarian and Tunisian strains preferred higher irradiances.     

CONCORDANCE OF MOLECULAR AND ULTRASTRUCTURAL DATA IN THE STUDY OF ZOOSPORIC CHLOROCOCCALEAN GREEN ALGAE1

Alternative evolutionary hypotheses generated from features of vegetative cell morphology and motile cell ultra-structure were investigated using a molecular data set. Complete nuclear-encoded small subunit (18S) ribosomal RNA (rRNA) gene sequences were determined for six species (three each) of the chlorococcalean green algae “Neo chloris” and Characium. Based on motile cell ultra-structure, it was previously shown that both genera could be separated into three distinct groups possibly representing three separate orders and two classes of green algae. 18S rRNA gene sequences were also obtained for three additional taxa, Dunaliella parva Lerche, Pediastrum duplex Meyen, and Friedmannia israelensis Chantanachat and Bold. These organisms were selected because each, in turn, is a representative of one of the three ultrastructural groups into which the Neochloris and Characium species are separable. Phylogenetic analyses utilizing the molecular data fully support the ultrastructural findings, suggesting that the similar vegetative cell morphologies observed in these organisms have resulted from convergence.     

Some rare desmids (Zygnematophyceae) from Central Europe

The morphology and taxonomy of eight rare desmids (Zygnematophyceae) from Central Europe are studied, i.e. six taxa from Slovakia [Tortitaenia bahusiensis (Nordstedt et Lütkemüller) Coesel, Closterium limneticum Lemmermann var. fallax R??i?ka, Euastrum sublobatum de Brébisson in Ralfs, morpha, Cosmarium simplicius (W. et G.S. West) Grönblad, Pachyphorium canadense (Irénée-Marie) G. H. Tomaszewicz et Hindák, stat. et comb. nova, Staurastrum bloklandiae Coesel et Joosten], one species from Poland (Spirotaenia bacillaris Lütkemüller) and Austria (Cosmarium geminatum Lundell). All desmids taxa are new records for the country of their origin.     

PHOTOSYNTHETIC CAPACITY AND LUXURY UPTAKE OF CARBON DURING PHOSPHATE LIMITATION IN PEDIASTRUM DUPLEX (CHLOROPHYCEAE)1

When cells of Pediastrum duplex Meyen experience phosphorus depletion, their capacity for carbon fixation declines, but sizes and carbon contents of the cells increase several-fold, an apparent instance of “luxury uptake” of carbon. Maximum rates of uptake of phosphate increase during the same period, and are consequently correlated with the enlarged surface area of the cells. Thus the disadvantage of increased cell volume, which may accelerate sinking speed in nature, is offset by the increased capacity of the cells for nutrient uptake.     

THE INFLUENCE OF pH ON ALGAL CELL MEMBRANE PERMEABILITY AND ITS IMPLICATIONS FOR THE UPTAKE OF LIPOPHILIC METAL COMPLEXES1

Uptake of lipophilic metal complexes by freshwater algae has recently been shown to be pH dependent. Here we look at different physiological aspects that could influence the diffusion of the lipophilic Cd complex, Cd(diethyldithiocarbamate)₂⁰ (Cd(DDC)₂⁰), into algal cells at different exposure pH values. Changes in cell membrane permeability were assessed as a function of pH for three species of green algae [Chlamydomonas reinhardtii P. A. Dang., Pseudokirchneriella subcapitata (Korshikov) Hindák, and Chlorella fusca var. vacuolata Shihira et R. W. Kraus] using two neutral, nonionic probes, fluorescein diacetate (FDA) and D‐sorbitol. In parallel experiments, we exposed algae to inorganic Cd or to Cd(DDC)₂⁰ and monitored Cd intracellular metal distribution, together with phytochelatin synthesis. For the three algal species acclimated at pH 5.5 (w/wo DDC 1 μM) and exposed at this pH, their permeability to FDA and D‐sorbitol was consistently lower than for algae growing at pH 7.0 and exposed at this pH (P < 0.001). The ratio of the FDA hydrolysis rate measured at pH 7.0 with respect to the rate measured at pH 5.5 (both in the presence of DDC) correlated with the ratio of the Cd(DDC)₂⁰ initial internalization rate constant obtained at pH 7.0 versus that obtained at pH 5.5 (three algae species, n = 9, r = 0.85, P = 0.004). Our results strongly suggest that acidification affects metal availability to algae not only by proton inhibition of facilitated metal uptake but also by affecting membrane permeability.     

Some new and uncommon British Volvocales,III

British material of the following taxa is described: Chlamydomonas mirabilis (Korshikov) Pascher var. minor nov. var., Chlamydomonas similis Korshikov var. curta nov. var., and Sphaerellopsis velata (Korshikov) Gerloff.     

Zoospore Structure and Colony Formation in Pediastrum Spp. and Hydrodictyon reticulatum (L.) Langerheim

The process of daughter colony formation in Pediastrum biradiatumMeyen, P. tetras (Ehren-berg) Ralfs, P. duplex Meyen, and Hydrodictyonreticulatum (L.) Lagerheim has been investigated using lightmicroscopy and electron microscopy of thin sections. The precisedisposition of the organelles confers upon the zoospores ofthese algae a special symmetry of a type unusual for the Chlorophyta.Apart from this, the detailed ultrastructure of the zoosporeand young vegetative cells resembles that of other green algaepreviously investigated. A common sequence of events has beenfound to occur during colony formation in both Pediastrum andHydrodictyon, with a correlation between zoospore symmetry andthat of the colony produced. The changes involved in the transitionfrom a motile to a non-motile cell are considered, and appearto have features in common with this process in some other greenalgae. The early stages in wall deposition involve the simultaneousformation of two distinct layers and serve to stick togetherthe cells which by this time have collectively assumed the basicmorphological characteristics of the mature coenobia.     

120 Exploration of Morphological Variation Within the Genus Pediastrum Meyen 1829 (Chlorophyceae, Chlorophyta)

Monographic works on the green algal genus, Pediastrum Meyen 1829 (Chlorophyceae, Chlorophyta), have described species, varieties and forms based on such characteristics as the size and shape of the marginal cells, pattern of cell wall sculpturing and extent of cell wall sculpturing. Depending on the author, the number of taxa assigned to the genus Pediastrum varies. Due to the lack of quantitative value to these characteristics, it has been difficult for other researchers to assign appropriate taxonomy to wild isolates. A molecular phylogeny including multiple strains from both culture collections and wild samples confirms relationships found by previous molecular studies on fewer taxa, in which the family Hydrodictyaceae forms a monophyletic group within the Sphaeropleales, and that the genera Hydrodictyon and Sorastrum are derived from Pediastrum . Hydrodicyton forms a monophyletic clade and consists of three species, H. reticulatum, H. africanum , and H. patenaeforme . Multiple isolates of H. reticulatum reveal little genetic variation between different geographic localities. Inclusion of wild isolates permits a more thorough exploration of morphological variation within the genus Pediastrum , and what characters may be taxonomically informative, particularly in the species P. boryanum and P. duplex . Wild isolates sampled from different areas also offers information regarding geographic variation and potential morphological convergence.     

Division of Chloroplasts in a Green Alga, Pediastrum duplex

The division of chloroplasts in a green alga, Pediastrum duplex,was studied by electron microscopy. Cells were treated for observationwith the freeze-substitution method. Fibrils, or fibrous belts,which we had observed previously at the dividing constrictionsof chloroplasts in Trebouxia potteri were not visible in Pediastrum,even though the method of preparation was the same for bothsets of samples. Microtubules (MTs) and the septum seem notto participate directly in the division of the single chloroplastin Pediastrum cells. Many thin fibrils, 7–20 nm in diameter,attached to, or protruding from, the surface of the dividingconstriction were seen. These fibres were less densely distributedat the constrictions of non-dividing chloroplasts. It is suggestedthat these fibrils are involved in the divison of chloroplastsin Pediastrum duplex. Cell wall, chloroplast division, freeze-substitution, intermediate fibres, Pediastrum duplex, transmission electron microscopy     

ULTRASTRUCTURE OF VEGETATIVE ORGANIZATION AND CELL DIVISION IN THE UNICELLULAR RED ALGA DIXONIELLA GRISEA GEN. NOV. (RHODOPHYTA) AND A CONSIDERATION OF THE GENUS RHODELLA1

This study suggests that the genus Rhodella be restricted to that set of features currently observed only in Rhodella maculata Evans and Rhodella violacea (Korn-mann) Wehrmeyer, that a new genus Dixoniella be established to accommodate the unicellular red alga, Rhodella grisea (Geitler) Fresnel, Billard, Hindák et Pekár-ková, and that Rhodella cyanea Billard et Fresnel be further studied for probable reclassification. These conclusions are based on ultrastructural comparisons of Dixoniella grisea with published information on the genus Rhodella. The presence of thylakoids in the pyrenoid, a peripheral encircling thylakoid in the chloroplast, a dictyosome/nuclear envelope association, and the lack of a specialized pyrenoid/nucleus association in D. grisea separate this alga from the genus Rhodella. Cell division in D. grisea is not demonstrably different from that in Rhodella, although the unusually well-defined material of the presumptive microtubule organizing center (MTOC) made it possible to follow the development and behavior of the MTOC to a greater degree than in previously studied red algal cells. The surprising amount of conformity in cell division characters between D. grisea and the genus Rhodella prompted a comparison of cell division characteristics in all red algal unicells studied to date. All unicells show a remarkable degree of similarity except for differences in interzonal spindle length, dissimilarities in size of the nucleus-associated organelle (NAO), and the unusual NAO of Porphyridium purpureum (Bory) Drew et Ross.     

Sensitivity of the Green Alga Pediastrum duplex Meyen to Allelochemicals Is Strain-Specific and Not Related to Co-Occurrence with Allelopathic Macrophytes

Interspecific differences in the response of microalgae to stress have numerous ecological implications. However, little is known of intraspecific sensitivities and the potential role of local genetic adaptation of populations. We compared the allelochemical sensitivity of 23 Pediastrum duplex Meyen strains, a common component of the freshwater phytoplankton. In order to test for local genetic adaptation, strains were isolated from water bodies with and without the allelopathically-active submerged macrophyte Myriophyllum. Strains were assigned to P. duplex on the basis of cell shape and colony morphology and only P. duplex strains that belonged to the same lineage in an ITS rDNA phylogeny were used. Inhibition of strain growth rates and maximum quantum yields of photosystem II were measured after exposure to tannic acid (TA) and co-culture with Myriophyllum spicatum. Growth rate inhibition varied over one order of magnitude between the P. duplex strains. There was no correlation between the presence of Myriophyllum in the source location and the sensitivity of the strains to TA or the presence of Myriophyllum, suggesting that at least strong unidirectional local adaptation to Myriophyllum had not taken place in the studied water bodies. The maximum quantum yield of photosystem II of TA exposed algae decreased, whereas the yield of algae exposed to M. spicatum was slightly higher than that of the controls. The ranking of P. duplex strain sensitivities differed between the types of exposure (single additions of TA versus co-existence with M. spicatum) and the parameter measured (growth rate versus maximum quantum yield), emphasizing the importance of measuring multiple traits when analysing strain-specific sensitivities towards allelochemicals. The observation that sensitivities to allelochemicals vary widely among strains of a single freshwater algal species should be taken into account if evaluating ecological consequences of allelopathic interactions.     

The Phylogenetic Relationships of the Saturn-shaped Ascospore-forming Species of the Genus Williopsis Zender and Related Genera Based on the Partial Sequences of 18S and 26S Ribosomal RNAs (Saccharomycetaceae): The Proposal of Komagataea Gen. Nov.

The partial base sequences of 18S and 26S rRNAs of strains of Williopsis and Saturnospora species were analyzed. In the three regions partially sequenced, the higher base differences were observed in the strains examined of the three species, W. californica, W. mucosa, and W. pratensis, compared with those of W. saturnus var. saturnus (type species of genus Williopsis), W. beijerinckii, W. mrakii, W. saturnus var. subsufficiens, W. suaveolens, P. membranaefaciens (type species of genus Pichia), C. matritensis (type species of genus Citeromyces), and S’spora dispora (type species of genus Saturnospora): the percent similarities were 52–82 in positions 493–622, 130 bases, of 26S rRNA, and the number of base differences was 28–6 in positions 1611–1835, 225 bases, of 26S rRNA, and the number of base differences was 25–4 in positions 1451–1618, 168 bases, of 18S rRNA. In the 18S rRNA partial base sequencings, W. mucosa had an identical base sequence with P. anomala (≡ H. anomala, type species of genus Hansenula). Based on the sequence data obtained, the taxonomic positions of the three Williopsis species mentioned above are discussed. The genus Zygowilliopsis Kudriavzev was postulated to be retained and emended, and a new genus, Komagataea was proposed for W. pratensis with a new combination, Komagataea pratensis.     

Protective Effect of Enzymatic Extracts from Microalgae Against DNA Damage Induced by H<Subscript>2</Subscript>O<Subscript>2</Subscript>

The enzymatic extracts from seven species of microalgae (Pediastrum duplex, Dactylococcopsis fascicularis, Halochlorococcum porphyrae, Oltmannsiellopsis unicellularis, Achnanthes longipes, Navicula sp. and Amphora coffeaeformis) collected from three habitats (freshwater, tidal pool, and coastal benthic) at Jeju Island in Korea were investigated for their antioxidant activity. Of the extracts tested, the AMG 300 L (an exo 1, 4-α-d-glucosidase) extract of P. duplex, the Viscozyme extract of Navicula sp., and the Celluclast extract of A. longipes provided the most potential as antioxidants. Meanwhile, the Termamyl extract of P. duplex in an H₂O₂ scavenging assay exhibited an approximate 60% scavenging effect. In this study, we report that the DNA damage inhibitory effects of P. duplex (Termamyl extract) and D. fascicularis (Kojizyme extract) were nearly 80% and 69% respectively at a concentration of 100 μg/ml. Thus, it is suggested that the microalgae tested in this study yield promising DNA damage inhibitory properties on mouse lymphoma L 5178 cells that are treated with H₂O₂. Therefore, microalgae such as P. duplex may be an excellent source of naturally occurring antioxidant compounds with potent DNA damage inhibition potential.     

Some new and uncommon British volvocales

British material of the following algae is described: Pyramimonas tetrarhynchus Schmarda; Spermatozopsis exsultans Korshikov; Myochloris collorhynchus nov. gen. et sp.; Pedinomonas minor Korshikov; Monomastix opisthostigma Scherffel; Lobomonas ampla Pascher var. okensis Korshikov; L. rostrata Hazen; Diplostauron pentagonium (Hazen) Pascher; Phacotus lendneri Chodat; Chlorobrachis sp.     

CORRESPONDENCE BETWEEN PHYLOGENY AND MORPHOLOGY OF SNOWELLA SPP. AND WORONICHINIA NAEGELIANA,CYANOBACTERIA COMMONLY OCCURRING IN LAKES1

In this study, the first reported isolates of the genera Snowella and Woronichinia were characterized by 16S rRNA gene sequencing and morphological analysis. Phylogenetic studies and sequences for these genera were not available previously. By botanical criteria, the five isolated strains were identified as Snowella litoralis (Häyrén) Komárek et Hindák Snowella rosea (Snow) Elenkin and Woronichinia naegeliana (Unger) Elenkin. This study underlines the identification of freshly isolated cultures, since the Snowella strains lost the colony structure and were not identifiable after extended laboratory cultivation. In the 16S rRNA gene analysis, the Snowella strains formed a monophyletic cluster, which was most closely related to the Woronichinia strain. Thus, our results show that the morphology of the genera Snowella and Woronichinia was in congruence with their phylogeny, and their phylogeny seems to support the traditional botanical classification of these genera. Furthermore, the genera Snowella and Woronichinia occurred commonly and might occasionally be the most abundant cyanobacterial taxa in mainly oligotrophic and mesotrophic Finnish lakes. Woronichinia occurred frequently and also formed blooms in eutrophic Czech reservoirs.     

UPTAKE OF LIPOPHILIC CADMIUM COMPLEXES BY THREE GREEN ALGAE: INFLUENCE OF HUMIC ACID AND ITS pH DEPENDENCE1

Cadmium forms neutral, lipophilic CdL₂⁰ complexes with diethyldithiocarbamate (L = DDC) and with ethylxanthate (L = XANT). In a synthetic solution and in the absence of natural dissolved organic matter (DOM), for a given total Cd concentration, uptake of these complexes by unicellular algae is much faster than the uptake of the free Cd²⁺ cation. The objective of the present study was to determine how this enhanced uptake of the lipophilic CdL₂⁰ complexes was affected by the presence of natural DOM (Suwannee River humic acid, SRHA). Experiments were performed with Cd(DDC)₂⁰ and Cd(XANT)₂⁰ at two pH values (7.0 and 5.5) and with the three chlorophytes [Chlamydomonas reinhardtii P. A. Dang., Pseudokirchneriella subcapitata (Korshikov) Hindák, Chlorella fusca var. vacuolata Shihira et R. W. Krauss]. Short‐term uptake (30–40 min) of the CdL₂⁰ complexes was followed in the absence and presence of SRHA (6.5 mg C · L^?1). Acidification from pH 7.0 to 5.5 decreased CdL₂⁰ uptake by the three algae, in the presence or absence of humic acid (HA). The dominant effect of the HA was to decrease Cd uptake, due to its interaction with the CdL₂⁰ complexes in solution. However, if uptake of the free CdL₂⁰ complexes was compared in the presence and absence of HA, in four of eight cases initial uptake rate constants (k_i) were significantly higher (P < 0.05) in the presence of the HA, suggesting the operation of an interfacial effect of the HA at the algal cell membrane, favoring uptake of CdL₂⁰. Overall, the experimental results suggest that neutral metal complexes will be less bioavailable in natural waters than they are in synthetic laboratory media in the absence of natural DOM.