Lobed Chloroplast Research Articles

Emendation of Rhodomonas marina (Cryptophyceae): insights from morphology, molecular phylogeny and water-soluble pigment in an Arctic isolate

Rhodomonas (Cryptophyceae) and species assigned to this genus have undergone numerous taxonomic revisions. This also applies to R. marina studied here as it was originally assigned as a species of Cryptomonas and later considered a variation of R. baltica, the type species. Despite being described more than 130 years ago, R. marina still lacks a comprehensive characterization. Light and electron microscopy were employed to delineate a strain from western Greenland. The living cells were 18 μm long and 9 μm wide, elliptical in shape with a pointed to rounded posterior and truncated anterior in lateral view. Two sub-equal flagella emerged from a vestibulum, where also a furrow extended. In transmission electron microscopy, the furrow was associated with a tubular gullet and the pyrenoid embedded in a deeply lobed chloroplast. The chloroplast contained DNA in perforations and was surrounded by starch grains. A tubular nucleomorph was enclosed within the pyrenoid matrix. In scanning electron microscopy, the inner periplast consisted of rectangular plates with rounded edges and posteriorly these were replaced by a sheet-like structure. The water-soluble pigment was Crypto-Phycoerythrin type I (Cr-PE 545). A phylogenetic inference based on SSU rDNA confirmed the identity of strain S18 as a species of Rhodomonas as it clustered with congeners but also Rhinomonas, Storeatula, and Pyrenomonas. These genera formed a monophyletic clade separated from a diverse assemblage of other cryptophyte genera. To further explore the phylogeny of R. marina a concatenated phylogenetic analysis based on the SSU rDNA-ITS1-5.8S rDNA-ITS2- LSU rDNA region was performed but included only closely related species. The secondary structure of nuclear internal transcribed spacer 2 was predicted and compared to similar structures in related species. Using morphological and molecular signatures as diagnostic features the description of R. marina was emended.

Chloromonas fuhrii sp. nov. (Chlorophyceae), a cosmopolitan alga from colored snow

ABSTRACT The modern concept of the genus Chloromonas includes a subclade of species found exclusively in snow. We describe Chloromonas (Cr.) fuhrii sp. nov., a new member of this clade, which is closely related to Cr. muramotoi from Japan. The new species is characterised by the absence of a stigma, pyrenoid, and defined papilla, ready loss of flagella in culture, and a single lobed chloroplast. Strains of Cr. fuhrii have been sourced from green, orange, and pink/red snow. The species has been found in Svalbard, Antarctica, and New Zealand, with most genetic variation occurring in Svalbard. The minimal variation present in New Zealand strains suggests a relatively recent arrival. Secondary structure models for ITS2 in the new species demonstrate a separation from Cr. muramotoi due to one CBC. The nine New Zealand strains isolated showed no variation in ITS2 but four variable base positions in ITS1. Inclusion of the single Antarctic and four Svalbard strains resulted in 14 variable positions in ITS2 and 69 in ITS1, with the sequences remaining alignable. The ITS data showed a strong biogeographic signal, both in total in an unrooted tree, and using ITS2 alone with Cr. muramotoi as an outgroup. The absence of a stigma in this species prompted a survey of the literature, showing that stigma absence in Chloromonas is strongly associated with the snow habitat, occurring at about 10 times the rate observed in other habitats, perhaps owing to the difficulty of phototaxis in snow with abundant reflected light.

Fine-structural characterization and phylogeny of Sphaerodinium (Suessiales, Dinophyceae), with the description of an unusual type of freshwater dinoflagellate cyst

Two strains of Sphaerodinium were established from two mountain areas in Portugal and examined by light microscopy, scanning and transmission electron microscopy, and sequence analyses of nuclear-encoded SSU, ITS1–5.8S–ITS2 and LSU rDNA. Both strains were identified as S. polonicum var. tatricum on the basis of comparison with the original taxonomic descriptions within the genus. The two strains were nearly identical in morphology and ultrastructure, except for the presence of pseudograna-like thylakoid stacks within more rounded chloroplast lobes in one of them. Sexual reproduction occurred in culture batches and resting cysts with single or grouped processes with wide bases and distal platforms with slightly recurved margins were seen to develop by sudden retraction of planozygote cytoplasm. Morphological, fine-structural and molecular characters were compared with previously available information from S. cracoviense, allowing for a more robust characterization of the genus. Important characters include a type F eyespot, a pusule canal linking the transverse flagellar canal to a collecting chamber connected to regular pusular tubes, a ventral fibre extending from the proximal-right side of the longitudinal basal body, and a membranous, lamellar body with a honeycomb pattern near the flagellar base area. The latter two features are shared with Baldinia anauniensis.

Biomineralization Capacities of Chlorodendrophyceae: Correlation Between Chloroplast Morphology and the Distribution of Micropearls in the Cell

Several species of the genus Tetraselmis (Chlorodendrophyceae, Chlorophyta) were recently discovered to possess unsuspected biomineralization capacities: they produce multiple intracellular inclusions of amorphous calcium carbonate (ACC), called micropearls. Early light-microscopists had spotted rows of refractive granules in some species, although without identifying their mineral nature. Scanning electron microscope (SEM) observations showed that the distribution of the micropearls in the cell forms a pattern, which appears to be characteristic for a given species. The present study shows that this pattern correlates with the shape of the chloroplast, which differs between Tetraselmis species, because micropearls align themselves along the incisions between chloroplast lobes. This was observed both by SEM and in live cells by light microscopy (LM) using Nomarski differential interference contrast. Additionally, molecular phylogenetic analyses, of rbcL and ITS2 gene sequences from diverse strains of Chlorodendrophyceae, corroborated the morphological observations by identifying two groups among nominal Tetraselmis spp. that differ in chloroplast morphology, micropearl arrangement, and ITS2 RNA secondary structure.

The golden paradox - a new heterokont lineage with chloroplasts surrounded by two membranes.

A marine, sand-dwelling, golden-brown alga is described from clonal cultures established from a high intertidal pool in southeastern Australia. This tiny, unicellular species, which we call the "golden paradox" (Chrysoparadoxa australica gen. et sp. nov.), is benthic, surrounded by a multilayered cell wall and attached to the substratum by a complex adhesive plug. Each vegetative cell gives rise to a single, naked zoospore with heterokont flagella that settles and may become briefly amoeboid prior to dividing. Daughter cells are initially amoeboid, then either permanently attach and return to the benthic stage or become motile again prior to final settlement. Two deeply lobed chloroplasts occupy opposite ends of the cell and are surrounded by only two membranes. The outer chloroplast membrane is continuous between the two chloroplasts via the outer membrane of the nuclear envelope. Only two membranes occupy the chloroplast-nucleus interface, the inner membrane of the nuclear envelope and the inner chloroplast membrane. A small pyrenoid is found in each chloroplast and closely abuts the nucleus or protrudes into it. It contains an unusual, membrane-bound inclusion that stains with SYBR green but is unlikely to be a nucleomorph. Phylogenies inferred from a 10-gene concatenated alignment show an early-branching position within the PX clade. The unusual morphological features and phylogenetic position indicate C.australica should be classified as a new class, Chrysoparadoxophyceae. Despite an atypical plastid, exploration of the C.australica transcriptome revealed typical heterokont protein targeting to the plastid.

Morphology and molecular phylogeny of Hyalosynedra lanceolata sp. nov. and an extended description of Hyalosynedra (Bacillariophyta)

ABSTRACTThe araphid diatom genus Hyalosynedra is a very common component of marine benthic communities, occurring as an epiphyte or colonizing hard substrata, and has a worldwide distribution. The hyaline appearance of the valve and the high striation density, which is indiscernible under a light microscope, makes the identification of most species difficult and very few studies have investigated their morphology, phylogeny or ecology in detail. In an extensive study of diatom communities from the hypersaline Mar Menor coastal lagoon (SE Spain), several new taxa were found. Herein we propose Hyalosynedra lanceolata sp. nov. based on morphological observations (light and electron microscopy) and molecular data (phylogenetic and sequence divergence analyses). The species has distinctive morphological characters (radiate colony, biseriate striation, lanceolate sternum and two long laminar lobed chloroplasts) that are new for the genus and have been included in a proposal to extend the genus description. Our results also show that H. toxoneides does not belong to Hyalosynedra, but appears to be more closely related to Thalassionema and Thalassiothrix. However, further molecular and morphological research is needed to clarify its taxonomic position.

Taxonomic revision and species delimitation of coccoid green algae currently assigned to the genus Dictyochloropsis (Trebouxiophyceae, Chlorophyta).

Coccoid green algae traditionally classified in Dictyochloropsis have a complex, reticulate chloroplast, when mature, without a pyrenoid. They occupy remarkably diverse ecological niches as free-living organisms or in association with lichen-forming fungi and were recently shown to form two distinct lineages within Trebouxiophyceae. We used a polyphasic approach to revise the taxonomy of the genus. Based on phylogenetic analysis of the 18S rRNA gene, and detailed morphological investigation using comparative conventional light and confocal microscopy, we have assigned these lineages to two genera, Dictyochloropsis and Symbiochloris gen. nov. We have reconsidered the diagnostic generic features as follows: Dictyochloropsis comprises only free-living algae with a reticulate chloroplast, forming lobes in a parallel arrangement at some ontogenetic stages, and which reproduce only by means of autospores. This agrees with Geitler's original diagnosis of Dictyochloropsis, but not with the later emendation by Tschermak-Woess. Consequently, the species of Dictyochloropsis sensu Tschermak-Woess are assigned to Symbiochloris, with new combinations proposed. Symbiochloris encompasses free-living and/or lichenized algae with lobed chloroplasts and that reproduce by forming zoospores characterized by two subapical isokont flagella that emerge symmetrically near the flattened apex. In addition, using coalescent-based approaches, morphological characters and secondary structure of ITS transcripts, we inferred species boundaries and taxonomic relationships within the newly proposed genera. Two species of Dictyochloropsis and nine species of Symbiochloris are delimited, including the newly described species D.asterochloroides, S.handae, S.tropica, and S.tschermakiae. Our results further support the non-monophyly of autosporine taxa within Trebouxiophyceae.

Andersenia, a genus of filamentous, sand-dwelling Pelagophyceae from southeastern Australia

:A new marine pelagophyte genus, Andersenia, was described from clonal cultures established from sand samples collected in coastal tide pools from southeastern Australia. Thin, uniseriate, unbranched filaments were variable in length up to 1.0 mm, 7–12 μm in width and surrounded by a gelatinous envelope. Cells contained one or two golden brown, lobed chloroplasts. Growing filaments often segmented into fragments of varying length that were continuous within the filament envelope and eventually produced multicellular knots, giving mature filaments a gnarled appearance. Most vegetative cells generated asexual motile zoospores that were heterokont with two unequal, heterodynamic flagella that arose together approximately one-third the length of the cell from the anterior surface; one or two lobed chloroplasts were located towards the posterior of the cells, while numerous granules of uniform size were typically obvious at the cell periphery. During segmentation, zoospore formation and release, the unusual cross walls shaped like biconcave discs were revealed and were characteristic of the genus. Zoospores adhered and secreted an adhesive pad within minutes of attachment and initiated new filaments. Phylogenetic analysis based on the nuclear-encoded small subunit ribosomal RNA (SSU rRNA) as well as morphological differences placed the new genus Andersenia in the class Pelagophyceae, and the three strains studied formed two species, A. nodulosa sp. nov. and A. australica sp. nov. Andersenia nodulosa filaments were longer and the cells smaller than in A. australica, and they differed by 16 nucleotides (99.2% identity) at the SSU rRNA sequence level.

Studies on woloszynskioid dinoflagellates VI: description of Tovellia aveirensis sp. nov. (Dinophyceae), a new species of Tovelliaceae with spiny cysts

A new species of Tovellia, T. aveirensis, is described on the basis of light (LM) and scanning electron microscopy (SEM) of motile cells and resting cysts, complemented with transmission electron microscopy (TEM) of flagellate cells and phylogenetic analysis of partial sequences of the large subunit ribosomal rRNA gene. Both vegetative cells and several stages of a life cycle involving sexual reproduction and the production of resting cysts were examined in cultures established from a tank in the University of Aveiro campus. Vegetative cells were round and little compressed dorsoventrally; planozygotes were longer and had a proportionally larger epicone. Chloroplast lobes were shown by TEM to radiate from a central, branched pyrenoid, although this was difficult to ascertain in LM. The amphiesma of flagellate cells had mainly 5 or 6-sided vesicles with thin plates, arranged in 5–7 latitudinal series on the epicone, 3–5 on the hypocone. The cingulum had 2 rows of plates, the posterior row extending into the hypocone and crossed by a series of small projecting knobs along the lower edge of the cingulum. A line of narrow amphiesmal plates extended over the cell apex, from near the cingulum on the ventral side to the middle of the dorsal side of the epicone. Eight or 9 narrow amphiesmal plates lined each side of this apical line of plates (ALP). Resting cysts differed from any described before in having numerous long, tapering spines with branched tips distributed over most of the surface. Most mature cysts showed an equatorial constriction. Neither cysts nor motile cells were seen to accumulate red cytoplasmic bodies in any stage of the cultures. The phylogenetic analysis placed, with high statistical support, the new species within the genus Tovellia; it formed a clade, with moderate support, with T. sanguinea, a species notable for its reddening cells.

Investigations of cell morphology and reproduction in Macrochloris radiosa Ettl & Gärtner (Stephano-sphaerinia, Chlorophyta) by light- and WUDQVPLVVLRQ electron microscopy.

Cell division and reproduction of a cultivated strain of Macrochloris radiosa (Stephanosphaerinia clade) were studied by light- and transmission electron microscopy. Multinucleate cells were frequently observed allowing description of the cell structure and details of the reproduction process. Nuclear staining revealed the position of the multiple polymorphic nuclei between the chloroplast lobes. Ultrastructure of coenocytic cells showed no signs of cleavage of the protoplast in the cytoplasm, although basal bodies were already present within the multinucleate cells. In the further course of the reproduction, biflagellate zoospores were developed that retained their flagella during sporulation. Zoospores were subsequently released from the sporangia.

TAXONOMIC REVISIONS OF MORPHOLOGICALLY SIMILAR SPECIES FROM TWO EUGLENOID GENERA: EUGLENA (E.GRANULATA AND E.VELATA) AND EUGLENARIA (EU.ANABAENA, EU.CAUDATA, AND EU.CLAVATA)(1).

The establishment of epitypes (together with the emended diagnoses) for three species of Euglenaria Karnkowska, E. W. Linton et Kwiatowski [Eu.anabaena (Mainx) Karnkowska et E. W. Linton; Eu.caudata (Hübner) Karnkowska et E. W. Linton; and Eu.clavata (Skuja) Karnkowska et E. W. Linton] and two species of Euglena Ehrenberg [E.granulata (Klebs) Schmitz and E.velata Klebs] was achieved due to literature studies, verification of morphological diagnostic features (cell size, cell shape, number of chloroplasts, the presence of mucocysts), as well as molecular characters (SSU rDNA). Now all these species are easy to identify and distinguish, despite their high morphological similarity, that is, spindle-shaped (or cylindrically spindle-shaped) cells and parietal, lobed chloroplasts with a single pyrenoid, accompanied by bilateral paramylon caps located on both sides of the chloroplast. E.granulata is the only species in this group that has spherical mucocysts. E.velata is distinguished by the largest cells (90-150 μm) and has the highest number of chloroplasts (>30). Eu.anabaena has the fewest chloroplasts (usually 3-6), and its cells are always (whether the organism is swimming or not) spindle-shaped or cylindrically spindle-shaped, in contrast to the cells of Eu.clavata, which are club-shaped (clavate) while swimming and only after stopping change to resemble the shape of a spindle or a cylindrical spindle; Eu.clavata has numerous chloroplasts (15-20). Eu.caudata is characterized by asymmetrical spindle-shaped (fusiform) cells, that is, with an elongated rear section and a shorter front section; the number of chloroplasts normally ranges from 7 to 15.

Chaetomorpha philippinensis (Cladophorales, Chlorophyta), a new marine microfilamentous green alga from tropical waters

Leliaert F., Payo D.A., Calumpong H.P. and De Clerck O. 2011. Chaetomorpha philippinensis (Cladophorales, Chlorophyta), a new marine microfilamentous green alga from tropical waters. Phycologia 50: 384–391. DOI: 10.2216/10-76.1A new marine microfilamentous green alga, Chaetomorpha philippinensis Leliaert sp. nov., is described as an epiphyte on Chaetomorpha vieillardii from shallow subtidal habitats in the Philippines. Phylogenetic analyses of large subunit rDNA and rDNA internal transcribed spacer (ITS) sequences show that the new tropical species is sister to the cold-water Chaetomorpha norvegica, from which it is genetically clearly distinct but morphologically almost indistinguishable. Chaetomorpha philippinensis is characterized by minute, straight or curved, unbranched, erect filaments up to 300 µm long and 7–17 µm in diameter, attached by a basal, hapteroid holdfast. Filaments in culture are similar in morphology but grow considerably longer with slightly larger cells. The cylindrical cells are multinucleate with up to eight nuclei (up to 18 in culture). Cells contain a single, parietal, lobed chloroplast with numerous small perforations and one to several pyrenoids. Zoosporangia develop by transformation of apical and subapical cells with zoids emerging through a domed pore in the apical, middle or basal part of the cell.

Mating system, auxosporulation, species taxonomy and evidence for homoploid evolution in Amphora (Bacillariophyta)

Mann D.G. and Poulíčková A. 2010. Mating system, auxosporulation, species taxonomy and evidence for homoploid evolution in Amphora (Bacillariophyta). Phycologia 49: 183–201. DOI: 10.2216/09-08.1Cytological characteristics of the mitotic cycle, sexual reproduction and auxospore formation are described for three members of the type group of Amphora: A. ovalis, A. copulata and A. minutissima. All have a single lobed ventral chloroplast, nuclei with granular heterochromatin and a single nucleolus (unusually large in A. ovalis). Amphora copulata is heterothallic, with two mating types that do not differ in gamete behaviour. All three species reproduce allogamously via fusion of two rearranged gametes per gametangium to produce elongate zygotes enclosed within the volume delimited by the gametangial thecae. Pairing is strictly via the ventral sides of the cells, with tight apposition of the cells, and expansion of the auxospores is strictly perpendicular to the gametangium long axes. Comparisons are made with other Amphora species, particularly A. proteus (also belonging to the subgenus Amphora) and A. arcus. Meiosis is described in detail in A. copulata, revealing no movement of the nucleus away from the dorsal position it occupies during the mitotic cycle, a synizetic knot in early prophase, no cytokinesis accompanying meiosis II, and rapid degeneration of the supernumerary nuclei in the gametic cells; the usual two acytokinetic mitoses accompany formation of the initial epivalve (which lacks a functional raphe) and hypovalve. The potential for formation of autopolyploids is demonstrated in A. copulata. Nevertheless, chromosome counts of 2n = 34 in A. copulata, 2n = 28 or 30 in A. ovalis and an undetermined but not significantly lower number in A. minutissima, suggest that homoploid evolution is more likely than polyploid. In mixed sexualized populations of the species, no interbreeding occurs, supporting separation of minutissima from copulata. Measurements of initial cell sizes, coupled with slight differences in valve width and pattern, suggest that A. copulata may be a species complex.

EVOLUTIONARY ORIGIN OF GLOEOMONAS (VOLVOCALES, CHLOROPHYCEAE), BASED ON ULTRASTRUCTURE OF CHLOROPLASTS AND MOLECULAR PHYLOGENY1

Gloeomonas is a peculiar unicellular volvocalean genus because it lacks pyrenoids in the chloroplasts under the light microscope and has two flagellar bases that are remote from each other. However, ultrastructural features of chloroplasts are very limited, and no molecular phylogenetic analyses have been carried out in Gloeomonas. In this study, we observed ultrastructural features of chloroplasts of three species of Gloeomonas and Chloromonas rubrifilum (Korshikov ex Pascher) Proschold, B. Marin, U. Schlosser et Melkonian SAG 3.85, and phylogenetic analyses were carried out based on the combined data set from 18S rRNA, ATP synthase beta-subunit, and P700 chl a-apoprotein A2 gene sequences to deduce the natural phylogenetic positions of the genus Gloeomonas. The present EM demonstrated that the chloroplasts of the three Gloeomonas species and C. rubrifilum SAG 3.85 did not have typical pyrenoids with associated starch grains, but they possessed pyrenoid matrices that protruded interiorly within the stroma regions of the chloroplast. The pyrenoid matrices were large and broad in C. rubrifilum, whereas those of the three Gloeomonas species were recognized in only the small protruded regions of the chloroplast lobes. The present multigene phylogenetic analyses resolved that the three species of Gloeomonas belong to the Chloromonas lineage or Chloromonadinia of the Volvocales, and Chloromonas insignis (Anakhin) Gerloff et H. Ettl NIES-447 and C. rubrifilum SAG 3.85, both of which have pyrenoids without associated starch grains, were positioned basally to the clade composed of the three species of Gloeomonas. Therefore, Gloeomonas might have evolved from such a Chloromonas species through reduction in pyrenoid matrix size within the chloroplast and by separating their two flagellar bases.

ULTRASTRUCTURE AND LSU rDNA-BASED REVISION OF PERIDINIUM GROUP PALATINUM (DINOPHYCEAE) WITH THE DESCRIPTION OF PALATINUS GEN. NOV.(1).

The name Peridinium palatinum Lauterborn currently designates a freshwater peridinioid with 13 epithecal and six cingular plates, and no apical pore complex. Freshwater dinoflagellate floras classify it in Peridinium group palatinum together with P.pseudolaeve M. Lefèvre. General ultrastructure, flagellar apparatus, and pusular components of P.palatinum were examined by serial section TEM and compared to P.cinctum (O. F. Müll.) Ehrenb. and Peridiniopsis borgei Lemmerm., respectively, types of Peridinium and Peridiniopsis. Partial LSU rDNA sequences from P.palatinum, P.pseudolaeve and several peridinioids, woloszynskioids, gymnodinioids, and other dinoflagellates were used for a phylogenetic analysis. General morphology and tabulation of taxa in group palatinum were characterized by SEM. Differences in plate numbers, affecting both the epitheca and the cingulum, combine with differences in plate ornamentation and a suite of internal cell features to suggest a generic-level distinction between Peridinium group palatinum and typical Peridinium. The branching pattern of the phylogenetic tree is compatible with this conclusion, although with low support from bootstrap values and posterior probabilities, as are sequence divergences estimated between species in group palatinum, and typical Peridinium and Peridiniopsis. Palatinus nov. gen. is proposed with the new combinations Palatinus apiculatus nov. comb. (type species; syn. Peridinium palatinum), P.apiculatus var. laevis nov. comb., and P.pseudolaevis nov. comb. Distinctive characters for Palatinus include a smooth or slightly granulate, but not areolate, plate surface, a large central pyrenoid penetrated by cytoplasmic channels and radiating into chloroplast lobes, and the presence of a peduncle-homologous microtubular strand. Palatinus cells exit the theca through the antapical-postcingular area.

Confocal microscopy of chloroplast morphology and ontogeny in three strains of Dictyochloropsis (Trebouxiophyceae, Chlorophyta)

P. Škaloud, J. Neustupa, B. Radochová and L. Kubínová. 2005. Confocal microscopy of chloroplast morphology and ontogeny in three strains of Dictyochloropsis (Trebouxiophyceae, Chlorophyta). Phycologia 44: 261–269.Chloroplast morphology and ontogeny in three species of the genus Dictyochloropsis – D. splendida var. splendida, D. reticulata and D. symbiontica – were investigated by using light and confocal microscopy. In a conventional light microscope, the complicated net-shaped chloroplast often appeared as a homogenous mass filling up most of the cell volume, while confocal microscopy enabled a detailed description of the chloroplast changes during its ontogeny. We identified four distinct morphological stages during the chloroplast ontogeny in all investigated strains. The stages are distinguished primarily by the number of differently structured chloroplast layers and by the inner structure of chloroplast lobes. The investigated Dictyochloropsis strains differed mainly in timing of these particular ontogenetic sequences. In the final stage of the chloroplast ontogeny, the transformation of the net-shaped chloroplast to a simple form allows the chloroplast division.

Taxonomic characterization of Pyramimonas aurea sp. nov. (Prasinophyceae, Chlorophyta)

An undescribed species of Pyramimonas, P. aurea, sp. nov. from the Republic of Palau has been examined by light and electron microscopy using cultured material. The cells have a typical pyramimonad shape with body and flagellar scales. The cells have numerous puncta along each chloroplast sinus and each anterior cleft of the chloroplast lobes, box scales with three spines on their side walls, crown scales with four uprights from the corners, limuloid scales with three subsidiary spines and five subsidiary radial ribs, segmented T-hair and Pt-hair scales and a 3–1 type flagellar apparatus. In addition, cells contain a single pyrenoid that is not invaded by thylakoids, and numerous centrally positioned vacuoles that contain fibrillar material. From these results, the alga belongs to the subgenus Punctatae and its closest relative is P. mucifera. The clear differences between it and P. mucifera are the cell dimensions and the morphology of the box and crown body scales. These differences clearly show that the alga is a new species of Pyramimonas. Phylogenetic analyses using 18S ribosomal DNA (rDNA) and rbcL sequences were obtained. The rbcL tree showed that each of the subgenera Pyramimonas sensu stricto, Trichocystis and Vestigifera is monophyletic and this is supported by relatively high bootstrap values. However, species of subgenus Punctatae, P. olivacea (the type species of the subgenus), P. formosa and P. aurea did not form a monophyletic clade. The heterogeneous nature of the subgenus Punctatae was also indicated in the 18S rDNA sequence analysis. These data suggest that P. formosa and P. aurea could not belong to the subgenus Punctatae and should be separated independently. However, further ultrastructural and molecular data are necessary to reconstruct the taxonomic system of the genus Pyramimonas.

STRUCTURE AND ASEXUAL REPRODUCTION OF THE ENIGMATIC CHAROPHYCEAN GREEN ALGA ENTRANSIA FIMBRIATA (KLEBSORMIDIALES, CHAROPHYCEAE)1

As the closest relatives of embryophytes, the charophycean green algae (sensu Mattox and Stewart) may reveal the evolutionary history of characters in this lineage. Recent molecular phylogenetic analysis indicates that the little‐known species Entransia fimbriata Hughes is a member of the charophycean order Klebsormidiales. In this study LM and EM were used to identify and describe additional structural characters of Entransia so that comparisons could be made with Klebsormidium and with other charophycean algae outside the order Klebsormidiales. Features that Entransia shares with various members of the genus Klebsormidium include cylindrical cells in unbranched filaments that may spiral, parietal chloroplasts that cover only part of the circumference of the cell, H‐shaped cross walls, and vegetative reproduction by both fragmentation and formation of zoospores or aplanospores. Among the characteristics that distinguish Entransia from Klebsormidium are a highly lobed chloroplast with multiple pyrenoids; a single large vacuole; short cells that die and collapse, apparently facilitating filament fragmentation; and germinating filaments with condensed adhesive at the base and a tapering spine at the tip. Although Entransia has sometimes been tentatively considered to be a member of the Zygnemataceae, the presence of a flagellate life history stage distinguishes Entransia from this group. The pyrenoids of Entransia are typical of those of charophycean algae in having traversing membranes and surrounding starch. Presence of multiple such pyrenoids in each chloroplast of Entransia supports the hypothesis that the common ancestor of charophycean algae and embryophytes had a single chloroplast with multiple pyrenoids.

Ultrastructural study of the type species of Peridiniopsis, Peridiniopsis borgei (Dinophyceae), with special reference to the peduncle and flagellar apparatus

The general ultrastructure and flagellar apparatus of Peridiniopsis borgei were studied by serial section electron microscopy. The cells contained a conspicuous multiple-stalked pyrenoid in an anterior-dorsal position, relative to a large, central sac pusule (SP), and were surrounded by an unusual mucilage layer, located between the thecal plates and the outer amphiesmal membranes. The central SP was linked to the longitudinal flagellar canal by a narrow, 1.5 μm long tubular extension. An apical fibrous complex with three radiating fibres was associated with the apical pore complex. A flat peduncle emerged near the flagellar bases and was supported by a single row of 75–80 microtubules, which extended for nearly 50 μm along an elaborate path inside the cell. The flagellar apparatus resembled that of Peridinium cinctum but differences were noted. The layered connective was smaller and had fewer thin layers than that of P. cinctum. The microtubular root associated with the anterior surface of the transverse basal body nucleated a novel type of transverse microtubular root extension, in which 23 microtubules surrounding a fibrouscore extended along the surface of the SP. Two distinct fibres connected triplets of the transverse basal body to the longitudinal microtubular root (LMR). A large eyespot, located within chloroplast lobes, was found in the sulcal area. It differed from previously described dinoflagellate eyespots by having up to six layers of electron-opaque globules; the inner layers were intercalated among thylakoid lamellae. A vesicle containing a layer of crystal-like bodies was present between the ventral surface of the eyespot and the LMR. In all cells sectioned, a double unit membrane was located between the thecal plates and the cytoplasmic membrane.

Chloroplast rotation and morphological plasticity of the unicellular alga Rhodosorus (Rhodophyta, Stylonematales)

SUMMARY Time-lapse videomicroscopy was used to film chloroplast rotation in the 13 isolates of Rhodosorus marinus Geitler and one isolate of Rhodosorus magnei Fresnel & Billard maintained in culture. Cell diameter, rates of chloroplast rotation and the number of chloroplast lobes were measured in all isolates. Chloroplast rotation is a definitive characteristic of the genus Rhodosorus and should be included in its taxonomic description. Isolates of the type species, R. marinus, measure 4–7 μm in diameter in low light (2–4 μmol photons m−2 s−1) and 4.5–11.5 μm in diameter in bright light (15–20 μmol photons m−2 s−1), and have two to seven chloroplast lobes. Rhodosorus magnei is 9 μm in diameter and has seven to nine chloroplast lobes in bright light. However, these cells are much smaller (approx. 4 μm diam.) and have only two to three chloroplast lobes when maintained in low light. The species R. magnei was created based on its larger cell size and numerous chloroplast lobes compared with R. marinus, but since these characteristics were found to be quite variable and dependant on culture conditions, they cannot be used to differentiate these two species.