Vladislav Cepák

Generic characters of the simplest cyanoprokaryotes Cyanobium, Cyanobacterium and Synechococcus

Abstract The cytomorphological features (cell morphology, type of cell division, cell structure, structure of photosynthetic apparatus) were studied in the type strains of the cyanoprokaryotic genera Cyanobium and Cyanobacterium , and in the reference strain of the traditional genus Synechococcus ( Synechococcus PCC 6301; = “ Anacystis nidulans ” sensu auct.). They were originally described by Rippka & Cohen-Bazire, based on the mean DNA-base composition (moles % G + C) and on the resistance to various cyanophages. The phenotypic diacritical characters were found to coincide well with the molecular markers, and thus, the genera Cyanobium and Cyanobacterium are acceptable also according to the traditional (botanical) taxonomic criteria. The integrated molecular, cytomorphological and ecophysiological approaches to the solution of taxonomic problems in cyanobacteria are therefore inevitable for taxonomic evaluation, from the methodological point of view. The lists of species within the revised genera Cyanobium and Cyanobacterium are reviewed.

The microalga Parachlorella kessleri––A novel highly efficient lipid producer

The alga Parachlorella kessleri, strain CCALA 255, grown under optimal conditions, is characterized by storage of energy in the form of starch rather than lipids. If grown in the complete medium, the cultures grew rapidly, producing large amounts of biomass in a relatively short time. The cells, however, contained negligible lipid reserves (1–10% of DW). Treatments inducing hyperproduction of storage lipids in P. kessleri biomass were described. The cultures were grown in the absence or fivefold decreased concentration of either nitrogen or phosphorus or sulfur. Limitation by all elements using fivefold or 10‐fold diluted mineral medium was also tested. Limitation with any macroelement (nitrogen, sulfur, or phosphorus) led to an increase in the amount of lipids; nitrogen limitation was the most effective. Diluted nutrient media (5‐ or 10‐fold) were identified as the best method to stimulate lipid overproduction (60% of DW). The strategy for lipid overproduction consists of the fast growth of P. kessleri culture grown in the complete medium to produce sufficient biomass (DW more than 10 g/L) followed by the dilution of nutrient medium to stop growth and cell division by limitation of all elements, leading to induction of lipid production and accumulation up to 60% DW. Cultivation conditions necessary for maximizing lipid content in P. kessleri biomass generated in a scale‐up solar open thin‐layer photobioreactor were described. Biotechnol. Bioeng. 2013; 110: 97–107.

Production of lipids and formation and mobilization of lipid bodies in Chlorella vulgaris

We investigated the formation of lipid bodies in the microalga Chlorella vulgaris CCALA 256 under lipid-induction conditions in autotrophically grown cultures. We found that cell division ceased after depletion of nitrates from the growth medium within the first days of cultivation. The growth of non-dividing cells subsequently led to the rapid accumulation of lipids. We describe in detail both biogenesis and mobilization of lipid bodies using fluorescence and transmission electron microscopy. Small lipid bodies fused very soon after their creation in the cytosol, forming, eventually, one huge lipid body. In stationary growth phase, lipids were present in the form of a large lipid body occupying most of the cell volume. After replenishment of nitrogen, lipid content decreased rapidly and, within 24 hours, the large lipid body was fragmented into smaller ones. This mobilization of the cellular lipid store occurred independently of light.

Cytoskeletal alterations in interphase cells of the green alga Spirogyra decimina in response to heavy metals exposure: I. The effect of cadmium

Summary.The aim of the study was to elucidate the effect of cadmium ions on the arrangement of the actin and tubulin cytoskeleton, as well as the relationships between cytoskeletal changes and growth processes in the green filamentous alga Spirogyra decimina. Batch cultures of algae were carried out under defined conditions in the presence of various cadmium concentrations. In control cells, the cytoskeleton appeared to be a transversely oriented pattern of both microtubules and actin filaments of various thickness in the cell cortex; colocalization of cortical microtubules and actin filaments was apparent. Microtubules were very sensitive to the presence of cadmium ions. Depending on the cadmium concentration and the time of exposure, microtubules disintegrated into short rod-shaped fragments or they completely disappeared. A steep increase in cell width and a decrease in growth rate accompanied (and probably ensued) a very rapid disintegration of microtubules. Actin filaments were more stable because they were disturbed several hours later than microtubules at any cadmium concentration used. When cadmium ions were washed out, the actin cytoskeleton was rebuilt even in cells in which actin filaments were completely disintegrated at higher cadmium concentrations (40 or 100 μM). The much more sensitive microtubules were regenerated after treatment with lower cadmium concentrations (10 or 15 μM) only.

ZOOSPOROGENESIS, MORPHOLOGY, ULTRASTRUCTURE, PIGMENT COMPOSITION, AND PHYLOGENETIC POSITION OF TRACHYDISCUS MINUTUS (EUSTIGMATOPHYCEAE, HETEROKONTOPHYTA)1

The traditional order Mischococcales (Xanthophyceae) is polyphyletic with some original members now classified in a separate class, Eustigmatophyceae. However, most mischococcalean species have not yet been studied in detail, raising the possibility that many of them still remain misplaced. We established an algal culture (strain CCALA 838) determined as one such species, Trachydiscus minutus (Bourr.) H. Ettl, and studied the morphology, ultrastructure, life cycle, pigment composition, and phylogeny using the 18S rRNA gene. We discovered a zoosporic part of the life cycle of this alga. Zoospore production was induced by darkness, suppressed by light, and was temperature dependent. The zoospores possessed one flagellum covered with mastigonemes and exhibited a basal swelling, but a stigma was missing. Ultrastructural investigations of vegetative cells revealed plastids lacking both a connection to the nuclear envelope and a girdle lamella. Moreover, we described biogenesis of oil bodies on the ultrastructural level. Photosynthetic pigments of T. minutus included as the major carotenoids violaxanthin and vaucheriaxanthin (ester); we detected no chl c. An 18S rRNA gene‐based phylogenetic analysis placed T. minutus in a clade with species of the genus Pseudostaurastrum and with Goniochloris sculpta Geitler, which form a sister branch to initially studied Eustigmatophyceae. In summary, our results are inconsistent with classifying T. minutus as a xanthophycean and indicate that it is a member of a novel deep lineage of the class Eustigmatophyceae.

Trachydiscus minutus, a new biotechnological source of eicosapentaenoic acid

The yellow-green alga Trachydiscus minutus (class Xanthophyta) was cultivated in a standard medium and in media without sulfur and nitrogen. Its yield after a 16-d cultivation reached 13 g dry mass per 1 L medium. The content of oligoenoic (‘polyenoic’) fatty acid (PUFA), i.e. eicosapentaenoic (EPA), was in excess of 35 % of total fatty acids; the productivity was thus 88 mg/L per d. This result makes the alga a very prospective organism that may serve as a new biotechnological source of single cell oil.

Cytoskeletal alterations in interphase cells of the green alga Spirogyra decimina in response to heavy metals exposure: II. The effect of aluminium, nickel and copper.

The effect of the toxic metal ions, aluminium (Al3+), nickel (Ni2+), and copper (Cu2+), on both the actin and tubulin cytoskeleton of the green alga Spirogyra decimina was studied. Batch cultured cells were grown for different time intervals at concentrations of 10, 15, 40 and 100 microM of aluminium as AlCl3, nickel as NiCl2 and copper as CuSO(4).5H2O. The impact of copper on the morphology of both MTs and AFs was much more prominent than the other two metals. A rapid irreversible depolymerization of cytoskeletal structures occurred, whereas in the presence of aluminium or nickel, changes in the cytoskeleton were slight and reversible to some extent. Nickel changed the orientation of cortical MTs, which turned from a transverse to a skewed or longitudinal direction. Aluminium caused slight depolymerization of the cytoskeleton, which reverted spontaneously to the normal cytoskeletal state (in AlCl3 free nutrient solution). Copper exerted a strong effect on both the MT and AF cytoskeleton, which fragmented and disorganized rapidly. The extent of cytoskeletal damage by copper was dosage and time dependent and AFs were slightly more sensitive than MTs.

What are the cyanobacterial genera Cyanothece and Cyanobacterium? Contribution to the combined molecular and phenotype taxonomic evaluation of cyanobacterial diversity

It is stressed the demand of the complex (molecular + biochemical + cytomorphological + ecological) evaluation of diversity of any taxonomic classification of cyanobacteria including coccal ones.

Cell Growth and Division Processes Are Differentially Sensitive to Cadmium in Scenedesmus quadricauda

The effect of cadmium on growth processes (accumulation of RNA, proteins and cell volume), cell cycle reproductive events (DNA replication, mitosis, protoplast fission and daughter-cell formation) and the regulatory activity of histone H1 kinases were monitored in synchronized cultures of the chlorococcal algaScenedesmus quadricauda. Distinct dosage-dependent inhibitory effects of cadmium ions were found in individual growth and reproductive processes. At concentration of about 60 µmol/L CdCl2, the growth processes were slowed down after about half of the cell cycle but the cells grew to the same or larger size than did untreated cells. At higher concentration, the growth became progressively inhibited, being completely blocked above 240 µmol/L. Total RNA accumulation was the most sensitive growth process. Each of the reproductive events was a target for cadmium ions with increasing sensitivity in the following order: DNA replication, mitosis, protoplast fission and daughter cell formation. Throughout the entire experiment, the activity of “mitosis-specific” histone H1 kinases was negligible in the cadmium (60 µmol/L CdCl2) treated cultures, whilst that of the control culture varied, peaking just prior to nuclear divisions. The activity of “growth-associated” histone H1 kinases was not affected by cadmium ions. No effect was found if cadmium was present during the precommitment period. The longer the period in the presence of cadmium, the stronger inhibition of reproductive events.

Cytomorphological characters supporting the taxonomic validity ofCyanothece (Cyanoprokaryota)

The fine structure of the type species of the genusCyanotheceKomárek 1976,C. aeruginosa, is described and compared with the main cytological characteristics of morphologically related members of the generaCyanobium, Cyanobacterium andSynechococcus. Several morphological features, such as cell walls with thick outer layers containing a special type of vesicles, position of thylakoids, “keritomy” (net-like appearance of protoplast caused by arrangement of thylakoids, net-like nucleoids and/or by tendency to form intrathylakoidal spaces) and a special structure of mucilaginous envelopes were found to be characteristic of this genus, supporting its separate position among coccal cyanoprokaryotes (cyanobacteria, cyanophytes). The taxonomic significance of ultrastructural features in all mentioned genera is discussed.