Dariusz Dziga

First report of the cyanobacterial toxin cylindrospermopsin in the shallow, eutrophic lakes of western Poland.

Cyanobacterial dominance in eutrophic lakes causes water quality problems due to the production of toxins harmful to humans and animals, as well as a number of odorous compounds. Cylindrospermopsin (CYN) is a potent cytotoxic cyanobacterial metabolite involved in triggering illness in humans. The occurrence of CYN has been mostly associated with tropical and subtropical cyanobacteria. We analyzed CYN concentration and phytoplankton assemblages of three lakes located in western Poland during the summers of 2006 and 2007. CYN was detected in 46% of our samples using the HPLC and LC-MS/MS methods. CYN concentrations were in the range of 0.16-1.8 microg L(-1) and exceeded the drinking water guideline value of 1 microgL(-1) in two samples. This is the first report of CYN occurrence in this part of Europe and provides further evidence that this toxin is common not only in subtropical and tropical regions. The lakes were dominated by Planktothrix agardhii but the occurrence of the CYN investigated here might be associated with the invasive species Cylindrospermopsis raciborskii and/or native Aphanizomenon gracile.

Microbial Degradation of Microcystins

Hepatotoxic microcystins that are produced by freshwater cyanobacteria pose a risk to public health. These compounds may be eliminated by enzymatic degradation. Here, we review the enzymatic pathways for the degradation of these hepatotoxins, some of which are newly discovered processes. The efficiencies of microcystin biodegradation pathways are documented in several papers and are compared here. Additionally, a comprehensive description of the microcystin enzymatic degradation scheme has been supplemented with a proposal for a new biodegradation pathway. Critical comments on less documented hypotheses are also included. The genetic aspects of biodegradation activity are discussed in detail. We also describe some methods that are useful for studying the biological decomposition of microcystins, including screening for microcystin degraders and detecting microcystin degradation products, with an emphasis on mass spectrometric methodology.

Carbohydrate and free amino acid contents in tomato plants grown in media with bicarbonate and nitrate or ammonium

Tomato plants were cultivated (from 2 to 23 days after germination) in media with NO3−, NH4+, or a mixture of both forms in different proportions used as the N source given with or without 5 mol dm−3 HCO3−. The accumulation of soluble sugars (reducing sugars and sucrose) and free amino acids was higher in the roots and leaves of NH4+-fed plants than in NO3−-fed plants. Starch accumulation in NH4+-fed plants was higher in leaves (about 28%) and lower in roots (about 37%) in comparison with that of NO3−-fed plants. Plants cultivated in media containing a mixture of NO3−/NH4+ were characterized by a lower content of sugars and amino acids accumulation in comparison with that in plants fed with NO3− or NH4+. An elevated HCO3− concentration in the rhizosphere stimulated the accumulation of soluble sugars and free amino acids in all the experimental variants. There were only small differences in the starch content.

Characterization of microcystin-LR removal process in the presence of probiotic bacteria.

Toxic cyanobacteria have been reported in lakes and reservoirs in several countries. The presence of toxins in drinking water creates a potential risk of toxin transference for water consumers. Besides chemical and physical methods of cyanotoxin removal from water, biodegradation methods would be useful. The aim of the current study was to identify bacterial removal mechanisms of the hepatotoxin microcystin-LR. This was studied by testing the hypothesis of enzymatic degradation of microcystin-LR in the presence of probiotic lactic acid bacterial and bifidobacterial strains and the participation of the proteolytic system of the bacteria in this process. The results suggest that extracellularly located cell-envelope proteinases are involved in the decomposition of microcystin-LR. In particular, a correlation between proteolytic activity and microcystin removal was found and both these parameters were dependent on glucose as an energy source. In addition, EDTA, which was indicated as a main inhibitor of proteinases of the investigated strain, was shown to limit the rate of microcystin removal. The removal of microcystins was shown to be different from the known microcystin-degradation pathway of Sphingomonas. (14)C-labeled microcystin was not found inside the cells and bacterial cell extracts were not able to remove the toxin, which supports the involvement of extracellularly located proteinases. The results confirm the hypothesis of enzymatic degradation of microcystins in the presence of probiotic bacteria.

Bioreactor study employing bacteria with enhanced activity toward cyanobacterial toxins microcystins.

An important aim of white (grey) biotechnology is bioremediation, where microbes are employed to remove unwanted chemicals. Microcystins (MCs) and other cyanobacterial toxins are not industrial or agricultural pollutants; however, their occurrence as a consequence of human activity and water reservoir eutrophication is regarded as anthropogenic. Microbial degradation of microcystins is suggested as an alternative to chemical and physical methods of their elimination. This paper describes a possible technique of the practical application of the biodegradation process. The idea relies on the utilization of bacteria with a significantly enhanced MC-degradation ability (in comparison with wild strains). The cells of an Escherichia coli laboratory strain expressing microcystinase (MlrA) responsible for the detoxification of MCs were immobilized in alginate beads. The degradation potency of the tested bioreactors was monitored by HPLC detection of linear microcystin LR (MC-LR) as the MlrA degradation product. An open system based on a column filled with alginate-entrapped cells was shown to operate more efficiently than a closed system (alginate beads shaken in a glass container). The maximal degradation rate calculated per one liter of carrier was 219.9 µg h−1 of degraded MC-LR. A comparison of the efficiency of the described system with other biological and chemo-physical proposals suggests that this new idea presents several advantages and is worth investigating in future studies.

Characterization of Enzymatic Activity of MlrB and MlrC Proteins Involved in Bacterial Degradation of Cyanotoxins Microcystins

Bacterial degradation of toxic microcystins produced by cyanobacteria is a common phenomenon. However, our understanding of the mechanisms of these processes is rudimentary. In this paper several novel discoveries regarding the action of the enzymes of the mlr cluster responsible for microcystin biodegradation are presented using recombinant proteins. In particular, the predicted active sites of the recombinant MlrB and MlrC were analyzed using functional enzymes and their inactive muteins. A new degradation intermediate, a hexapeptide derived from linearized microcystins by MlrC, was discovered. Furthermore, the involvement of MlrA and MlrB in further degradation of the hexapeptides was confirmed and a corrected biochemical pathway of microcystin biodegradation has been proposed.

Fruit yield of tomato cultivated on media with bicarbonate and nitrate/ammonium as the nitrogen source

ABSTRACT This article presents the effects of nitrate/ammonium (NO3 −/NH4 +), applied at different proportions to the root media with or without 5 mmol bicarbonate (HCO3 −), on the yield and chemical composition of tomato fruit. Tomato plants were grown hydroponically (pH 6.9) in glasshouse conditions. The yield of fruit fresh matter from four clusters obtained from plants grown on the medium with NH4 + was about 25% lower than from the plants grown on the medium containing NO3 − as the nitrogen (N) source. Supplying NO3 −/NH4 + at a ratio of 4:1 increased the fruit yield by about 20% in comparison with the value recorded for NO3 −−plants. The enrichment of the medium with HCO3 − stimulated the bearing, while the result depended on the ratio of NO3 −/NH4 +. A combined treatment of HCO3 − with NO3 − or NH4 + in the medium increased yields by about 28% and 11%, respectively, in comparison to plants cultivated without HCO3 −. The application of NO3 −/NH4 + at ratios of 4:1 and 1:1 with HCO3 − increased the respective yields by about 16% and 10% in comparison with plants grown without HCO3 −. Modifications in the composition of the media affected the accumulation of organic solutions in the fruit. The NH4 + nutrition effected a 20% decrease in the accumulation of reducing sugars in the fruit in comparison to the fruit of plants grown in media with NO3 −. In the cultivation of plants in media with various NO3 −/NH4 + proportions the intermediate values of the reduced sugar concentrations were recorded in comparison with the values obtained for NO3 −−plants and NH4 +−plants. The enrichment of media with HCO3 − increased the concentration of sugars in fruit from about 28% (for NO3 −−plants) to about 10% (for NH4 +−plants). Malate and citrate are the main constituents of carboxylates in tomato fruit. The form of nitrogen applied to the medium did not significantly affect the concentration of carboxylates in fruit. Significant differences in carboxylate concentrations appeared in fruit grown on media enriched with HCO3 − ions. In comparison with the cultivation without HCO3 −, increases in the accumulation of carboxylates varied from about 22% to 30% depending on the form of the applied nitrogen. The concentration of amino acids in the fruit of plants grown with NH4 + exceeded that in NO3 −−plants by about 55%. In the plants grown on media of modified NO3 −/NH4 + proportions, the concentration of amino acids in fruits were positively correlated with the level of NH4 + in the medium. The enrichment of media with HCO3 − stimulated a further increase in amino acid concentration in fruit by about 9% in NO3 − plants and about 21% in NH4 + plants compared with the respective control (without HCO3 −).

Biochemical and Morphological Alterations in Rat Liver Golgi Complexes After Treatment with Bis(maltolato)oxovanadium(IV) [BMOV] orMaltol Alone

Summary Oral treatment with maltol or bis(maltolato)oxovanadi- um(IV) [BMOV] alters the biochemical activity of the rat liver Golgi marker enzyme, i.e., galactosyltrans- ferase (GalT), and the organelle morphology in a relatively short time. Four groups of rats were investigated: control (C), treated with BMOV for 2 days (pVC), treated with BMOV for 7 days (C+V), and treated with maltol alone for 7 days (C+M). All drugs were administered as drinking solutions. These conditions were used, because normalization of galactosyltransferase activity (GalT) and morphology of rat liver Golgi complexes were previously found by us in streptozotocin-induced diabetes. In this paper, we present the influence of BMOV or maltol alone (as a vanadium ligand in BMOV compound) on rat liver Golgi complexes. The lowest statistically significant enzyme activity, in comparison with three other groups of rats (p

Cylindrospermopsin Biodegradation Abilities of Aeromonas sp. Isolated from Rusałka Lake

The occurrence of the cyanobacterial toxin cylindrospermopsin (CYN) in freshwater reservoirs is a common phenomenon. However, the biodegradation of this toxin in environmental samples has been observed only occasionally. In this work the biodegradation ability of cylindrospermopsin was investigated based on isolates from lakes with previous cyanotoxin history. Bacterial strains were identified based on the 16S rDNA and rpoD gene comparison. CYN biodegradation was monitored using the HPLC method. The R6 strain identified as Aeromonas sp. was documented as being capable of CYN removal. This biodegradation was dependent on the pH and temperature. Additionally, the stimulation of the growth of the R6 strain in the presence of CYN was indicated. Our discovery supports the hypothesis that (in analogy to the well-known phenomenon of microcystin biodegradation) in lakes dominated by potential CYN-producing cyanobacteria, the processes of microbial utilization of this toxin may occur.

EXTRACELLULAR ENZYMES OF THE MICROCYSTIS AERUGINOSA PCC 7813 STRAIN ARE INHIBITED IN THE PRESENCE OF HYDROQUINONE AND PYROGALLOL, ALLELOCHEMICALS PRODUCED BY AQUATIC PLANTS

Several cyanobacterial species have a high potential to dominate in marine environments and freshwater reservoirs, and the ecological and physiological reasons for this phenomenon are not understood comprehensively. In this study, the ability of a Microcystis aeruginosa Kütz. strain to produce free dissolved enzymes was documented. We have observed that this highly toxic strain releases alkaline phosphatase, leucine aminopeptidase, and β‐glucosidase into the ambient environment. Additionally, the inhibitory activity of selected phenols produced by aquatic plants on the activity of these enzymes was analyzed. The investigated compounds, pyrogallol and, to a lesser degree, hydroquinone, decreased the activity of extracellular enzymes produced by M. aeruginosa, with leucine aminopeptidase being the most sensitive to the inhibitors. The noncompetitive character of enzymatic inhibition suggests that the polyphenols produced by aquatic plants are able to influence the activity of different extracellular or membrane‐bound enzymes.