György Borbély

Comparative study of cyanotoxins affecting cytoskeletal and chromatin structures in CHO-K1 cells

In this study we compared the effects of the two frequently occuring and most dangerous cyanobacterial toxins on the cellular organization of microfilaments, microtubules and on the chromatin structure in Chinese hamster ovary (CHO-K1) cells. These compounds are the widely known microcystin-LR (MC-LR) and cylindrospermopsin (CYN) classified as the highest-priority cyanotoxin. Toxic effects were tested in a concentration and time dependent manner. The hepatotoxic MC-LR did not cause significant cytotoxicity on CHO-K1 cells under 20 microM, but caused apoptotic changes at higher concentrations. Apoptotic shrinkage was associated with the shortening and loss of actin filaments and with a concentration dependent depolymerization of microtubules. No necrosis was observed over the concentration range (1-50 microM MC-LR) tested. Cylindrospermopsin did cause apoptosis at low concentrations (1-2 microM) and over short exposure periods (12h). Necrosis was observed at higher concentrations (5-10 microM) and following longer exposure periods (24 or 48h). Cyanotoxins also affected the chromatin structure. The condensation process was inhibited by MC-LR at a later stage and manifested as broken elongated prechromosomes. CYN inhibited chromatin condensation at the early fibrillary stage leading to blurred fluorescent images of apoptotic bodies and preventing the formation of metaphase chromosomes. Cylindrospermopsin exhibited a more pronounced toxic effect causing cytoskeletal and nuclear changes as well as apoptotic and necrotic alterations.

Deep chlorophyll maximum by Ceratium hirundinella (O. F. Müller) Bergh in a shallow oxbow in Hungary

The stability of the water column permitted stable stratification of the shallow (maximum depth: 3.75 m) oxbow, Kecskészugi-Holt Körös in summer 2000. During the stratified period a deep chlorophyll maximum (DCM) was found at depths getting 60–180 μmol m−2 s−1 photosynthetically active radiation (PAR). The phytoplankton was dominated by Ceratium hirundinella and it is concluded that the development of the DCM largely resulted from the behavioural aggregation of this motile flagellate.

Cylindrospermopsin and microcystin-LR alter the growth, development and peroxidase enzyme activity of white mustard (Sinapis alba L.) seedlings, a comparative analysis

This work focuses on the comparative analysis of the effects of two cyanobacterial toxins of different chemical structure cylindrospermopsin (CYN) and microcystin-LR (MC-LR) on the white mustard (Sinapis alba L.) seedlings. Both cyanotoxins reduced significantly the fresh mass and the length of cotyledons, hypocotyls and main roots of seedlings in a concentration dependent manner. For various mustard organs the 50% inhibitory concentration values (IC50) of growth were between 3-5 μg ml(-1) for MC-LR and between 5-10 μg ml-1 for CYN, respectively. Cyanotoxins altered the development of cotyledons, the accumulation of photosynthetically active pigments and anthocyanins. Low MC-LR concentrations (0.01 and 0.1 μg ml(-1)) stimulated anthocyanin formation in the cotyledons but higher than 1 μg ml(-1) MC-LR concentrations strongly inhibited it. The CYN treated chlorotic cotyledons were violet coloured in consequence of high level of anthocyanins, while MC-LR induced chlorosis was accompanied by the appearance of necrotic patches. Necrosis and increases of peroxidase enzyme activity (POD) are general stress responses but these alterations were characteristic only for MC-LR treated mustard plants. These findings provide experimental evidences of developmental alterations induced by protein synthesis and protein phosphatase inhibitory cyanotoxins (CYN and MC-LR) in a model dicotyledonous plant.

Isolation of viable cell mass from frozen Microcystis viridis bloom containing microcystin-RR.

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.

Histological, cytological and biochemical alterations induced by microcystin-LR and cylindrospermopsin in white mustard (Sinapis alba L.) seedlings

This study compares the histological, cytological and biochemical effects of the cyanobacterial toxins microcystin-LR (MCY-LR) and cylindrospermopsin (CYN) in white mustard (Sinapis alba L.) seedlings, with special regard to the developing root system. Cyanotoxins induced different alterations, indicating their different specific biochemical activities. MCY-LR stimulated mitosis of root tip meristematic cells at lower concentrations (1 μg ml-1) and inhibited it at higher concentrations, while CYN had only inhibitory effects. Low CYN concentrations (0.01 μg ml-1) stimulated lateral root formation, whereas low MCY-LR concentrations increased only the number of lateral root primordia. Both inhibited lateral root development at higher concentrations. They induced lignifications, abnormal cell swelling and inhibited xylem differentiation in roots and shoots. MCY-LR and CYN induced the disruption of metaphase and anaphase spindles, causing altered cell divisions. Similar alterations could be related to decreased protein phosphatase (PP1 and PP2A) activities in shoots and roots. However, in vitro phosphatase assay with purified PP1 catalytic subunit proved that CYN in contrast to MCY-LR, decreased phosphatase activities of mustard in a non-specific way. This study intends to contribute to the understanding of the mechanisms of toxic effects of a protein phosphatase (MCY-LR) and a protein synthesis (CYN) inhibitory cyanotoxin in vascular plants.

Cylindrospermopsin inhibits growth and modulates protease activity in the aquatic plants Lemna minor L. and Wolffia arrhiza (L.) Horkel

The toxic effects of cylindrospermopsin (cyanobacterial toxin) on animals have been examined extensively, but little research has focused on their effects on plants. In this study cylindrospermopsin (CYN) caused alterations of growth, soluble protein content and protease enzyme activity were studied on two aquatic plants Lemna minor and Wolffia arrhiza in short-term (5 days) experiments. For the treatments we used CYN containing crude extracts of Aphanizomenon ovalisporum (BGSD-423) and purified CYN as well. The maximal inhibitory effects on fresh weight of L. minor and W. arrhiza caused by crude extract were 60% and 54%, respectively, while the maximum inhibitory effects were 30% and 43% in the case of purified CYN at 20 μg ml(-1) CYN content of culture medium. In CYN-treated plants the concentration of soluble protein showed mild increases, especially in W. arrhiza. Protease isoenzyme activity gels showed significant alterations of enzyme activities under the influence of CYN. Several isoenzymes were far more active and new ones appeared in CYN-treated plants. Treatments with cyanobacterial crude extract caused stronger effects than the purified cyanobacterial toxins used in equivalent CYN concentrations.

The pyridine nucleotide-dependent D-glucose dehydrogenase of Nostoc sp. strain Mac, a cyanobacterium, is subject to thioredoxin modulation

A pyridine nucleotide‐dependent D‐glucose dehydrogenase (GDH) was isolated and purified about 1000‐fold from Nostoc sp. strain Mac. The activity of this preparation with NADP as cofactor was 2.8‐times that with NAD. This ratio did not change during purification. The enzyme both in crude extracts and after purification proved to be subject to redox modulation. Homologous and heterologous (Anacystics nidulans, Anabaena sp. strain PCC 7120, spinach) thioredoxins, in the presence of 0.5 mM DTT, deactivated the enzyme. The thioredoxin from Nostoc was active with heterologous enzymes: it activated the fructose‐1,6‐bisphosphatase of Anacystis nidulans and the NADP‐dependent malate dehydrogenase of spinach. The thioredoxin‐mediated reduction decreased the apparent V max value for D‐glucose by about 65% and that for NADP by about 51%. The apparent K m value for NADP increased upon reduction by about 10‐fold. The apparent K m value for D‐glucose was but slightly affected by the redox state of the enzyme.

Seasonal succession of phytoplankton in a small oligotrophic oxbow and some consideration to the PEG model

Most of the research on the structure and dynamics of phytoplankton in lacustrine environments was performed in larger lakes. In small lakes, especially oxbows in which there is a profusion of waterbodies, available information is scarce (REYNOLDS 1993). Phytoplankton seasonal succession is discussed by comparison with the world model for lakes, PEG (Plankton Ecology Group, SoMMER et al. 1986). This model consists of 24 sequential statements, which describe, step by step, the seasonal events, which occur in phytoplankton and zooplankton of an idealised standard lake (Lake Constance). These statements have been tested using data from 24 different lakes, none of which were oxbows. The objectives of this paper are to describe the phytoplankton structure and to discuss the main patterns of seasonal variations of the phytoplankton species of a Hungarian oxbow. The present article is the first concerning an oxbow phytoplankton community and discusses data in relation to the steps proposed by the PEG model

Investigation of toxin content in Cylindrospermopsis raciborski (Wołoszyńska) Seenaya and Subba Raju and Aphanizomenon ovalisporum (Forti) strains isolated from shallow lakes of Hungary

Cylindrospermopsin (CYN) is an alkaloid type cytotoxic metabolite produced by several cyanobacterial species, which caused human illnesses. The occurrence of CYN has been mostly associated with tropical and subtropical cyanobacteria, but recently it is appearing in several countries, all over the world. We analyzed CYN concentration and polyketide synthase/peptide synthetase (PKS /PS) genes, important parts of the gene cluster responsible for the CYN biosynthesis, in 14 isolated/collected Cylindrospermopsis raciborskii and Aphanizomenon ovalisporum strains originated mostly from Hungary. CYN and PKS /PS genes were detected in Aphanizomenon ovalisporum strains isolated from Spain (of our isolation) and isolated in Israel (IL C-164), but the Hungarian isolate from the hyposaline Lake Szelidi had a lack of production capacity. In the Hungarian samples of C. raciborskii, we found no CYN and PKS /PS genes content comparing to CYN producer C. raciborskii AQS originated from Australia.

AS- 1 cyanophage infection inhibits the photosynthetic electron flow of photosystem II in Synechococcus sp. PCC 6301, a cyanobacterium

In Synechococcus sp. cells AS‐1 cyanophage infection gradually inhibits the photosystem II mediated photosynthetic electron flow whereas the activity of photosystem I is apparently unaffected by the cyanophage infection. Transient fluorescence induction and flash‐induced delayed luminescence decay studies revealed that the inhibition may occur at the level of the secondary acceptor, QB of photosystem II. In addition, the breakdown of D1‐protein is inhibited, comparable to DCMU‐induced protection of D1‐protein turnover, in AS‐1‐infected cells.