In the present study we provide cytological and biochemical evidence that the cyanotoxin cylindrospermopsin (CYN) induces programmed cell death (PCD) symptoms in two model vascular plants: the dicot ...white mustard (
Sinapis alba
) and the monocot common reed (
Phragmites australis
). Cytological data include chromatin fragmentation and the increase of the ratio of TUNEL-positive cells in roots, the latter being detected in both model systems studied. The strongest biochemical evidence is the elevation of the activity of several single-stranded DNA preferring nucleases-among them enzymes active at both acidic and alkaline conditions and are probably directly related to DNA breaks occurring at the initial stages of plant PCD: 80 kDa nucleases and a 26 kDa nuclease, both having dual (single- and double-stranded nucleic acid) specificity. Moreover, the total protease activity and in particular, a 53–56 kDa alkaline protease activity increases. This protease could be inhibited by PMSF, thus regarded as serine protease. Serine proteases are detected in all organs of
Brassicacea
e (Arabidopsis) having importance in differentiation of specialized plant tissue through PCD, in protein degradation/processing during early germination and defense mechanisms induced by a variety of biotic and abiotic stresses. However, knowledge of the physiological roles of these proteases and nucleases in PCD still needs further research. It is concluded that CYN treatment induces chromatin fragmentation and PCD in plant cells by activating specific nucleases and proteases. CYN is proposed to be a suitable molecule to study the mechanism of plant apoptosis.
Saffron is an expensive spice, cultivated in many regions of the world. Its chief metabolites include crocins, which are responsible for the coloring ability, safranal, which is the main essential ...oil constituent, and picrocrocin which is the main bitter constituent of the spice. A simple micellar capillary electrochromatographic (MEKC) method capable of quantifying all three types of main constituents was established. The pH, sodium dodecyl sulphate (SDS) content and electrolyte concentration of the background electrolyte was optimized. A simple extraction protocol was developed which can extract all metabolites of different polarity from the saffron stigmas. Optimal background electrolyte composed of 20mM disodium phosphate, 5mM sodium tetraborate, 100mM SDS, pH was set 9.5. Optimal extracting solvent was the background electrolyte, incubated with the sample for 60min. The proposed method allows quantification of picrocrocin, safranal, crocetin- Di-(β-d-gentiobiosyl) ester and crocetin (β-d-glycosyl)-(β-d-gentiobiosyl) ester within 17.5min, with limit of detection values ranging from 0.006 to 0.04mg/ml, from a single stigma.
Microcystin-LR (MC-LR) is a heptapeptide cyanotoxin, known to be a potent inhibitor of type 1 and 2A protein phosphatases in eukaryotes. Our aim was to investigate the effect of MC-LR on the ...organization of microtubules and mitotic chromatin in relation to its possible effects on cell and whole organ morphology in roots of common reed (
Phragmites australis).
P. australis is a widespread freshwater and brackish water aquatic macrophyte, frequently exposed to phytotoxins in eutrophic waters. Reed plantlets regenerated from embryogenic calli were treated with 0.001–40
μg
ml
−1 (0.001–40.2
μM) MC-LR for 2–20 days. At 0.5
μg
ml
−1 MC-LR and at higher cyanotoxin concentrations, the inhibition of protein phosphatase activity by MC-LR induced alterations in reed root growth and morphology, including abnormal lateral root development and the radial swelling of cells in the elongation zone of primary and lateral roots. Both short-term (2–5 days) and long-term (10–20 days) of cyanotoxin treatment induced microtubule disruption in meristems and in the elongation and differentiation zones. Microtubule disruption was accompanied by root cell shape alteration. At concentrations of 0.5–5
μg
ml
−1, MC-LR increased mitotic index at long-term exposure and induced the increase of the percentage of meristematic cells in prophase as well as telophase and cytokinesis of late mitosis. High cyanotoxin concentrations (10–40
μg
ml
−1) inhibited mitosis at as short as 2 days of exposure. The alteration of microtubule organization was observed in mitotic cells at all exposure periods studied, at cyanotoxin concentrations of 0.5–40
μg
ml
−1. MC-LR induced spindle anomalies at the metaphase–anaphase transition, the formation of asymmetric anaphase spindles and abnormal sister chromatid separation.
This paper reports for the first time that MC-LR induces cytoskeletal changes that lead to alterations of root architecture and development in common reed and generally, in plant cells. The MC-LR induced alterations in cells of an ecologically important aquatic macrophyte can reveal the importance of the effects of a cyanobacterial toxin in aquatic ecosystems.
The aim of this study was to establish the histological effects of exposure to microcystin-LR (MC-LR), a cyanotoxin, on axenic Phragmites australis plantlets. Plantlets were regenerated from ...embryogenic reed calli by tissue culture methods. Microcystin-LR inhibited the growth and development of embryogenic calli and the growth of reed plantlets. The 50% plantlet growth inhibitory concentration value (IC₅₀) of MC-LR was 12 μg ml⁻¹ (12.07 μ m) on mineral medium and 36 μg ml⁻¹ (36.22 μ m) on Murashige-Skoog medium. In the case of roots, the IC₅₀ value was 4.1 μg ml⁻¹ (4.12 μ m) on both media. Microcystin-LR induced aerenchyma obturation, altered lignification of cell walls in the axial organs, root necrosis and the capture of lateral or adventitious roots in the tissues of axial organs of reed plantlets. Cyanotoxin induced the premature development of lateral roots, root coalescence and early aerenchyma formation. Our data suggest that microcystin-LR, a cyanotoxin, induced developmental and histological alterations leading to growth inhibition of reed, and the induced harms have an impact on understanding reed decay in eutrophic fresh waters.
We aimed to study the histological and cytological alterations induced by cylindrospermopsin (CYN), a protein synthesis inhibitory cyanotoxin in roots of common reed (
Phragmites australis). Reed is ...an ecologically important emergent aquatic macrophyte, a model for studying cyanotoxin effects. We analyzed the histology and cytology of reed roots originated from tissue cultures and treated with 0.5–40
μg
ml
−1 (1.2–96.4
μM) CYN. The cyanotoxin decreased root elongation at significantly lower concentrations than the elongation of shoots. As general stress responses of plants to phytotoxins, CYN increased root number and induced the formation of a callus-like tissue and necrosis in root cortex. Callus-like root cortex consisted of radially swollen cells that correlated with the reorientation of microtubules (MTs) and the decrease of MT density in the elongation zone. Concomitantly, the cyanotoxin did not decrease, rather it increased the amount of β-tubulin in reed plantlets. CYN caused the formation of double preprophase bands; the disruption of mitotic spindles led to incomplete sister chromatid separation and disrupted phragmoplasts in root tip meristems. This work shows that CYN alters reed growth and anatomy through the alteration of MT organization.
Cyanobacterial species commonly occur in the phytoplankton of freshwater lakes and sometimes develop as toxin-producing blooms. Microcystis is one of the most common genera of freshwater ...cyanobacteria and is often the dominating phytoplankton of eutrophic lakes all over the world. In eutrophic lakes, large amounts of Microcystis may overwinter in the sediment and re-inoculate the water column in spring. In most cases, the overwintering pelagic population—if it exists—is small, and its role in re-inoculation has not been clear yet. In December 2005, we found large amounts of Microcystis on the surface, frozen in the ice cover in a eutrophic pond (Pond Hármashegy, Hungary). We identified the Microcystis species and investigated the viability and the toxicity of the frozen cells. The dominant species in the bloom samples was Microcystis viridis. Viability tests showed that the colonies isolated from the ice cover were composed of living cells. The isolated strain was found toxic, we analyzed the microcystin composition in the frozen planktonic Microcystis mass; in the investigated samples microcystin-RR was the main cyanotoxin.
► The morphogenesis and silicon response of
P. australis embryogenic callus cultures were studied. ► Plant regeneration potential depended on the genotype of explants. ► Silicon stimulated the growth ...of both stem nodal and root calli in a concentration dependent manner. ► Silicon stimulated somatic embryogenesis of root calli. ► Silicon influenced morphogenesis in a genotype-dependent manner.
The effects of silicon on the growth and development of
Phragmites australis (Cav.) Trin. Ex Steud. (common reed) stem nodal and root embryogenic calli were investigated. Silicon is considered to be a beneficial or quasi-essential nutrient for several Gramineaceous plants, including reed. Seven callus lines of four geographical locations (genotypes 1–4) within Hungary were investigated. Callus lines 1A, 2A and 3A were produced from stem nodal explants, while lines 1B, 2B, 3B and 4 were produced from roots. For the assay of silicon-dependent growth of callus lines of identical genotype but originating from different explants, we measured the increase of fresh weight of lines 1A and 1B. The studied developmental parameters were the increase of the number of somatic embryos (for callus lines 1A and 1B) and plant or root production from somatic embryos (for all genotypes/callus lines). Silicon was added to the culture medium as sodium silicate. In control cultures, plant or root regeneration from embryogenic calli was strongly genotype- and explant type-dependent. Stem nodal explants developed plants on regeneration medium in case of callus lines 2A and 3A, while line 1A produced roots only. All root derived calli developed roots on regeneration medium. Silicon stimulated the growth of both stem nodal and root calli (callus lines 1A, B) however, the concentration optima were different. Somatic embryogenesis of root calli, but not of stem nodal calli, was stimulated by silicate at low concentrations. However, for both of these callus lines, root development was stimulated by silicon. It had genotype-dependent influences on plant regeneration: while stimulation was observed in case of callus line 2A, inhibition occurred for line 3A. Root morphogenesis on calli was significantly influenced by silicon and depended on the callus line studied. Root production was stimulated on callus lines 1A, B and 2B, while in case of callus line 3B, it was significantly inhibited. The morphogenetic effects of Si were similar for different explants of the same geographical origin, i.e. plant or root production was similarly stimulated or inhibited by this element. We can conclude that the effects of Si on plant or root development depend on reed genotype used for callus induction. Its effect on growth and somatic embryogenesis depends on the explant type used for callus production. This is the first detailed report on the role of silicon in plant vegetative development and morphogenesis of a Gramineaceous plant.
Abstract
The effect of sulfate and phosphate deprivation on cell growth and cylindrospermopsin level was studied in Aphanizomenon ovalisporum ILC-164. Sulfate starvation induced a characteristic ...reduction of cylindrospermopsin pool size on the basis of cell number and unit of dry mass of culture. Phosphorous starvation of A. ovalisporum cultures induced a lesser reduction of cylindrospermopsin pool size. This divergence in the pool size of cylindrospermopsin may be the consequence of different growth rate. To show the metabolic changes concomitant with reduction of cylindrospermopsin pool size were obtained by measurement of ATP sulfurylase and alkaline phosphatase activity. The present study is the first concerning the cylindrospermopsin content under sulfate starvation and discusses it in relation to phosphorous starvation.
Capillary zone electrophoresis (CZE) and micellar electrokinetic chromatography (MEKC) were applied to the simultaneous separation of cyanobacterial toxins (anatoxin‐a, microcystin‐LR, ...cylindrospermopsin). The analytical performance data of both methods, optimized for the three toxins, were similar with a precision of migration times smaller than 0.8 RSD% and a detection limit in the range of 1–4 μg/mL, using spectrophotometric detection at 230 nm. Both methods were applied to an analysis of cyanotoxins in water bloom samples and crude cyanobacterial extracts. The results obtained indicate that, for complex matrices, the sequential application of CZE and MEKC is necessary. It is recommended to use both CE techniques for the analysis of the same sample in order to confirm the results by an orthogonal approach.
In the summer of 2006 bloom-like phenomenon occurred in a garden pond in Szeged, Southern Hungary. After regular watering of a sward with pond water containing the algal mass, destruction of garden ...grass occurred.
Microcystis aeruginosa, Microcystis viridis, Microcystis ichthyoblabe,
and
Microcystis wesenbergii
were identified by light microscopy in the water sample; microcystin-FR, -LR, -RR and -YR were determined by matrix-assisted laser desorption/ionization—time-of-flight analysis. There was an 80% decrease in the green mass (87% in chlorophyll-content) of the grass in a 1 m
2
area of the garden irrigated with pond water.
PDF
