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Toxicity of microcystins in the isolated hepatocytes of common carp (Cyprinus carpio L.).

The toxicity of hepatotoxic microcystins produced mainly by Microcystis aeruginosa in mammals and fishes was well studied in recent years. However, there were scarcely reports in toxic effects of microcystins on isolated hepatocytes of fishes, especially investigation of microcystin-induced apoptosis and/or necrosis in carp hepatocytes. In the present study, the isolated hepatocytes of common carp were exposed to various concentrations of microcystins (0.01, 0.1, 1, 10, 100, 1000 microg L(-1)) for 2, 4, 8, 16 and 24h, respectively, and cytotoxicity of microcystins in the toxin-treated cells was determined. Results of this study showed that cytotoxicity of microcystins on carp hepatocytes was time and dose-dependent, and the approximate LC(50) of microcystins in carp hepatocytes was 169.2 microg L(-1). The morphological changes typical of apoptosis, such as blebbing of cell membrane, condensation and fragmentation of cell nucleus were observed in the hepatocytes exposed to microcystins (1, 10 and 100 microg L(-1)) using fluorescence and differential interference contrast microscopy. Agarose gel electrophoresis of DNA demonstrated a typical apoptotic "ladder pattern" in microcystin-treated hepatocytes after 16 h of exposure. Results of the present study indicated that the form of cell death in microcystin-treated hepatocytes depend on the exposure dose of toxin. When lower concentration of microcystins (10 and 100 microg L(-1)) was used for exposure, carp hepatocytes died in apoptosis while, when higher one used (1000 microg L(-1)), they died in the form of necrosis.

Animals↗

A simple colorimetric method to detect biological evidence of human exposure to microcystins.

Toxic cyanobacteria are contaminants of surface waters worldwide. Microcystins are some of the most commonly detected cyanotoxins. Biological evidence of human exposure may be difficult to obtain due to limitations associated with cost, laboratory capacity, analytic support, and expertise. We investigated the application of an enzyme-linked immunosorbant assay (ELISA) to detect microcystins in human serum. We analyzed ten serum samples collected from dialysis patients who were known to be exposed to a mixture of microcystins during a 1996 outbreak in Brazil. We applied a commercially available ELISA method to detect microcystins in serum, and we compared the ELISA results to a more specific method, liquid chromatography/mass spectrometry (LC/MS) that was also used to detect microcystins in serum. The Spearman correlation coefficient was calculated using serum microcystin concentrations in split samples obtained by the two methods. Serum microcystin concentrations were similar, and we found good correlation of microcystin concentrations between the two methods. The ELISA detected total microcystins, median=19.9 ng/ml; LC/MS detected microcystin-LR equivalents, median=21.2 ng/ml; Spearman r=0.96, p<0.0001. We found that ELISA is a simple, accessible method to screen human serum for evidence of microcystin exposure.

Bacterial Toxins↗

Three new microcystins, cyclic heptapeptide hepatotoxins, from Nostoc sp. strain 152.

Three new cyclic heptapeptide hepatotoxins, [D-Ser1,ADMAdda5]microcystin-LR (1), [D-Asp3,-ADMAdda5]microcystin-LHar (2), and [ADMAdda5,Mser7]microcystin-LR (3), were isolated from the cyanobacterium (blue-green alga) Nostoc sp. strain 152, together with four known microcystins, [ADMAdda5]microcystin-LR (4), [ADMAdda5]microcystin-LHar (5), [D-Asp3,-ADMAdda5]microcystin-LR (6), and [DMAdda5]microcystin-LR (7). The structures of new microcystins were assigned on the basis of high-resolution fast atom bombardment mass spectrometry (HR FABMS), collisionally induced tandem FABMS (FABMS/MS), amino acid analysis, and gas chromatography (GC) on a chiral capillary column. All three new toxins contained 9-acetoxy-3-amino-2,6,8-trimethyl-10-phenyldeca-4,6-dienoic acid (ADMAdda) instead of the corresponding 9-methoxyl derivative (Adda), while 7 contains the corresponding 9-hydroxy analog (DMAdda). Compound 1 is the first microcystin reported that contains D-serine (D-Ser) in lieu of the D-alanine (D-Ala) unit which was thought to be an invariable amino acid component of the microcystins. Compound 2 has L-homoarginine (Har) instead of L-arginine (L-Arg) in 6 and D-aspartic acid (D-Asp) instead of D-erythro-beta-methylaspartic acid (D-MeAsp) in 5. Compound 3, the N-methylserine (Mser) variant of the N-methyldehydroalanine unit in 4, would be a biosynthetic precursor of 4.

Amino Acids↗

Seasonal variation and indirect monitoring of microcystin concentrations in Daechung reservoir, Korea.

Physicochemical and biological water quality, including the microcystin concentration, was investigated from spring to autumn 1999 in the Daechung Reservoir, Korea. The dominant genus in the cyanobacterial blooming season was Microcystis. The microcystin concentration in particulate form increased dramatically from August up to a level of 200 ng liter(-1) in early October and thereafter tended to decrease. The microcystin concentration in dissolved form was about 28% of that of the particulate form. The microcystins detected using a protein phosphatase (PP) inhibition assay were highly correlated with those microcystins detected by a high-performance liquid chromatograph (r = 0.973; P < 0.01). Therefore, the effectiveness of a PP inhibition assay for microcystin detection in a high number of water samples was confirmed as easy, quick, and convenient. The microcystin concentration was highly correlated with the phytoplankton number (r = 0.650; P < 0.01) and chlorophyll-a concentration (r = 0.591; P < 0.01). When the microcystin concentration exceeded about 100 ng liter(-1), the ratio of particulate to dissolved total nitrogen (TN) or total phosphorus (TP) converged at a value of 0.6. Furthermore, the microcystin concentration was lower than 50 ng liter(-1) at a particulate N/P ratio below 8, whereas the microcystin concentration varied quite substantially from 50 to 240 ng liter(-1) at a particulate N/P ratio of >8. Therefore, it seems that the microcystin concentration in water can be estimated and indirectly monitored by analyzing the following: the phytoplankton number and chlorophyll-a concentration, the ratio of the particulate and the dissolved forms of N and P, and the particulate N/P ratio when the dominant genus is toxigenic Microcystis.

Chlorophyll↗

Effects of light on the microcystin content of Microcystis strain PCC 7806.

Many cyanobacteria produce microcystins, hepatotoxic cyclic heptapeptides that can affect animals and humans. The effects of photosynthetically active radiation (PAR) on microcystin production by Microcystis strain PCC 7806 were studied in continuous cultures. Microcystis strain PCC 7806 was grown under PAR intensities between 10 and 403 micro mol of photons m(-2) s(-1) on a light-dark rhythm of 12 h -12 h. The microcystin concentration per cell, per unit biovolume and protein, was estimated under steady-state and transient-state conditions and on a diurnal timescale. The cellular microcystin content varied between 34.5 and 81.4 fg cell(-1) and was significantly positively correlated with growth rate under PAR-limited growth but not under PAR-saturated growth. Microcystin production and PAR showed a significant positive correlation under PAR-limited growth and a significant negative correlation under PAR-saturated growth. The microcystin concentration, as a ratio with respect to biovolume and protein, correlated neither with growth rate nor with PAR. Adaptation of microcystin production to a higher irradiance during transient states lasted for 5 days. During the period of illumination at a PAR of 10 and 40 micro mol of photons m(-2) s(-1), the intracellular microcystin content increased to values 10 to 20% higher than those at the end of the dark period. Extracellular (dissolved) microcystin concentrations were 20 times higher at 40 micro mol of photons m(-2) s(-1) than at 10 micro mol of photons m(-2) s(-1) and did not change significantly during the light-dark cycles at both irradiances. In summary, our results showed a positive effect of PAR on microcystin production and content of Microcystis strain PCC 7806 up to the point where the maximum growth rate is reached, while at higher irradiances the microcystin production is inhibited.

Bacterial Toxins↗

[Total analysis system for tumor promoter microcystins produced by cyanobacteria].

Microcystins, produced by freshwater cyanobacteria, are cyclic peptide hepatotoxins and tumor promoters. An outbreak of human poisoning attributed to microcystins has been reported in Caruaru, Brazil in 1996, where exposure through renal dialysis led to the death of 50 patients. Although such severe acute effects on human health seem to be rare, microcystins poses problems to human health which could result from low-level, chronic exposure to microcystins in drinking water. It is therefore important to monitor the levels of microcystins in water reservoirs where cyanobacterial blooms occur. We have developed a total analysis system for microcystins using GC-MS and LC-MS. This comprises initial screening of samples to check for the presence of microcystins by detecting 2-methyl-3-methoxy-4-phenylbutyric acid as an ozonolysis product using thermospray interface LC-MS and electron ionization/GC-MS. If a sample is positive in a screening test, it will be necessary to follow through with identification and quantification. Frit-FAB interface LC-MS allowed the rapid identification of microcystins in cyanobacteria and lake water, and also enabled us to identify microcystins and their metabolites formed in vivo in mouse liver. Finally, Frit-FAB/LC-MS using selected ion monitoring could be used for quantitative analysis of microcystins in lake water in the low nanogram range. The total analysis system proposed in the present study should be applicable to studies of the metabolism of microcystins, of their detoxification, and those of the mechanism(s) of the accumulation in the food chain.

Animals↗

Seven new microcystins possessing two L-glutamic acid units, isolated from Anabaena sp. strain 186.

Electrospray ionization mass spectrometry has been applied to the structure assignment of seven new microcystins (1-7), obtained from cultured Anabaena sp. strain 186. The seven new microcystins contain the dehydroalanine (Dha) or L-Ser unit instead of the N-methyldehydroalanine unit and the L-Glu and/or its delta-methyl ester [E(OMe)] units at the two variable L-amino acid units, and the structures were assigned as [Dha7]microcystin-E(OMe)E(OMe) (1), [D-Asp3,Dha7]microcystin-E(OMe)E(OMe) (2), [L-Ser7]microcystin-E(OMe)E(OMe) (3), [D-Asp3,L-Ser7]microcystin-E(OMe)E(OMe) (4), [Dha7]microcystin-EE(OMe) (5), [D-Asp3,Dha7]microcystin-EE(OMe) (6), and [L-Ser7]microcystin-EE(OMe) (7). These microcystins are the first examples containing dicarboxylic amino acids at the two variable L-amino acid units in microcystins.

Anabaena↗

Microcystin content of Microcystis aeruginosa is modulated by nitrogen uptake rate relative to specific growth rate or carbon fixation rate.

Modulation of microcystin production has been extensively studied in both batch and continuous cultures. Positive correlations with medium nitrogen, medium phosphorous, light intensity, inorganic carbon availability, and growth rate have been reported. Negative correlations have been reported between microcystin content and medium phosphorous. The only reported quantitative relationship between any variable and microcystin production was that of growth rate. Microcystis aeruginosa PCC7806 was therefore cultured under continuous culture conditions in a bubble-lift reactor at a growth rate of 0.01 h(-1) in modified BG11 (constant phosphate concentration of 0.195 mM and varying nitrate from 0.125 to 18 mM) and sampled at steady states for analysis of cell number, microcystin content, cellular N and P, residual medium nutrient concentration, and carbon fixation rate. Cellular microcystin quotas showed significant positive correlation with both nitrate uptake and cellular nitrogen content and were negatively correlated with carbon fixation rate, phosphate uptake, and cellular phosphorous. Thus, the ratio of nitrate uptake to phosphate uptake, cellular N to cellular P, and nitrate uptake to carbon fixation were positively correlated to cellular microcystin. Microcystin quotas increased 10-fold from the lowest to the highest steady-state values. Cellular microcystin content therefore is controlled to a significant extent by variables other than growth rate, as was previously reported, with nitrogen the most significant modulator. Batch culture in BG11 under identical conditions yielded increased microcystin when nitrogen uptake exceeded relative growth rate, confirming the importance of nitrogen uptake in the modulation of microcystin content for a specific growth rate.

Carbon↗

Microcystins induce morphological and physiological changes in selected representative phytoplanktons.

Dissolved microcystins (MC) are regularly present in water dominated by microcystin-producing, bloom-forming cyanobacteria. In vitro experiments with environmentally feasible concentrations (5 x 10(-7) M) of the three most common microcystins, MC-LR, -RR, and -YR, revealed that they influence the metabolism of different representative phytoplanktons. At light intensities close to the cyanobacterial bloom environment (50 mumol m(-2) s(-1)), they produce morphological and physiological changes in both microcystin-producing and nonproducing Microcystis aeruginosa strains, and also have similar effects on the green alga Scenedesmus quadricauda that is frequently present in cyanobacterial blooms. All three microcystin variants tested induce cell aggregation, increase in cell volume, and overproduction of photosynthetic pigments. All three effects appear to be related to each other, but are not necessarily caused by the same mechanism. The biological activity of microcystins toward the light-harvesting complex of photobionts can be interpreted as a signal announcing the worsening of light conditions due to the massive proliferation of cyanobacteria. Although the function of microcystins is still unknown, it is evident that they have numerous effects on phytoplankton organisms in nature. These effects depend on the individual organism as well as on the various intracellular and extracellular signaling pathways. The fact that dissolved microcystins also influence the physiology of microcystin-producing cyanobacteria leads us to the conclusion that the role of microcystins in the producing cells differs from their role in the water environment.

Eutrophication↗

Variations in the microcystin production of Planktothrix rubescens (cyanobacteria) assessed from a four-year survey of Lac du Bourget (France) and from laboratory experiments.

Between 1999 and 2002, a routine survey of water quality in the Lac du Bourget was performed to study the dynamics and microcystin (MC) production of Planktothrix rubescens. Using liquid chromatography coupled to diode array detection and mass spectrometry, we found that two main variants ([D-Asp3] and [D-Asp3, Dhb7] microcystin-RR) were produced. The proportion of these two variants was not influenced by the depth or season of sampling. Expressed in microcystin-LR equivalents, high microcystin concentrations were recorded from August to December each year, reaching values of up to 6.7 microg L-1. A significant correlation was found between the microcystin cell content and the cell densities of P. rubescens. Cellular quotas of microcystins ranged from 0.1 to 0.3 pg cell-1. Simultaneously, laboratory experiments were performed on a strain of P. rubescens isolated from the lake to assess the potential impact of various P-PO4 (3-) concentrations on intra- and extracellular microcystin production. Unlike natural populations, this strain only produced [D-Asp3] MC-RR. The intracellular microcystin content was similarly correlated to the cell density, but the cellular quota was slightly higher (0.3-0.7 pg cell-1) than in the natural population. Again, as in the natural population, a linear relationship was found between growth rate and microcystin production rate. These findings support the hypothesis that environmental factors, such as phosphate concentrations, have no direct impact on microcystin production by P. rubescens, but act indirectly by affecting growth rate.

Bacterial Toxins↗

Microcystins induce morphological and physiological changes in selected representative phytoplanktons.

Dissolved microcystins (MCs) are regularly present in water dominated by microcystin-producing, bloom-forming cyanobacteria. In vitro experiments with environmentally feasible concentrations (5 x 10(-7) M) of the three most common microcystins, MC-LR, MC-RR, and MC-YR, revealed that they influence the metabolism of different representative phytoplanktons. At light intensities that are close to the cyanobacterial bloom environment (50 micromol m(-2) s(-1)), they produce morphological and physiological changes in both microcystin-producing and -nonproducing Microcystis aeruginosa strains and also have similar effects on the green alga Scenedesmus quadricauda that is frequently present in cyanobacterial blooms. All three microcystin variants tested induce cell aggregation, increase in cell volume, and overproduction of photosynthetic pigments. All three effects appear to be related to each other but are not necessarily caused by the same mechanism. The biological activity of microcystins toward the light-harvesting complex of photobionts can be interpreted as a signal announcing the worsening of light conditions due to the massive proliferation of cyanobacteria. Although the function of microcystins is still unknown, it is evident that they have numerous effects on phytoplankton in nature. These effects depend on the individual organism as well as on the various intracellular and extracellular signaling pathways. The fact that dissolved microcystins also influence the physiology of microcystin-producing cyanobacteria leads us to the conclusion that the role of microcystins in the producing cells differs from the role in the water environment.

Fluorescence↗

Two new L-serine variants of microcystins-LR and -RR from Anabaena sp. strains 202 A1 and 202 A2.

Two new microcystins, [L-Ser7]microcystin-LR (1) and [L-Ser7]microcystin-RR (2), were isolated from a filamentous fresh water cyanobacterium (blue-green alga), Anabaena sp. strain 202 A1, along with the two major toxins, [Dha7]microcystin-LR (3) and [Dha7]microcystin-RR (4) and their minor components the D-Asp variants [D-Asp3,Dha7]microcystin-LR (5) and [D-Asp3,Dha7]microcystin-RR (6). Anabaena sp. strain 202 A1 also produced another new toxin, whose structure is tentatively proposed as [D-Asp3,L-Ser7]microcystin-XR (7), where X is a leucine homologue. Anabaena sp. strain 202 A2 produced one new microcystin, 1, and three known microcystins, 3, 4, and 5. The structures of the toxins were assigned based on their amino acid analyses, and fast atom bombardment mass spectrometry data.

Amino Acid Sequence↗

Identification and characterization of hydrophobic microcystins in Canadian freshwater cyanobacteria.

Hepatotoxic microcystins produced by cyanobacteria in freshwater lakes represent a significant health hazard to humans and agricultural livestock. Liquid chromatography (LC)-linked protein phosphatase (PPase) bioassay analysis of blooms of Microcystis aeruginosa produced in a Canadian drinking water lake identified several PPase inhibitors with significantly greater hydrophobicity than microcystin-LR, based on their retention time on C18 reverse phase LC columns. Seven PPase inhibitors were purified to homogeneity by bioassay-guided fractionation involving Sephadex LH-20 chromatography and two-step reverse phase at pH 6.5 and 2.0. One of the PPase inhibitors, isolated in a final yield of 1.5 micrograms/g lyophilized cyanobacteria, was identified as microcystin-LL by amino acid analysis and mass spectrometry. A further PPase inhibitor (20 ng/g cyanobacteria) was identified as microcystin-LL but with D-Ala replaced by an unknown amino acid. Four PPase inhibitors (< 20 ng/g cyanobacteria) were characterized by amino acid analysis and identified as microcystin-LV, -LM, -LF and -LZ (where Z represents an unknown hydrophobic amino acid). A further microcystin was also identified (< 10 ng/g cyanobacteria) in which arginine was apparently absent. The biological activity of the seven microcystins as inhibitors of the catalytic subunit of protein phosphatase-1 (PP-1c) was compared with microcystin-LR and motuporin (a hydrophobic analogue of nodularin). All of the compounds inhibited PP-1c with IC50 values of 0.06-0.4 nM, consistent with their identification as microcystins. These findings further demonstrate the applicability of a sensitive PPase bioassay for the identification of variant microcystins in the natural environment.

Amino Acids↗

Inhibition of several protein phosphatases by a non-covalently interacting microcystin and a novel cyanobacterial peptide, nostocyclin.

Microcystins produced by cyanobacterial 'blooms' in reservoirs and lakes pose significant public health problems because they are highly toxic due to potent inhibition of protein serine/threonine phosphatases in the PPP family. A dehydrobutyrine (Dhb)-containing microcystin variant [Asp3, ADMAdda5, Dhb7]microcystin-HtyR isolated from Nostoc sp. was found to potently inhibit PP1, PP2A, PPP4 and PPP5 with IC50 values similar to those of microcystin-LR. However, in contrast to microcystin-LR, which forms a covalent bond with a cysteine residue in these protein phosphatases, Asp,ADMAdda,Dhb-microcystin-HtyR did not form any covalent interaction with PP2A. Since the LD50 for Asp,ADMAdda,Dhb-microcystin-HtyR was 100 microg kg(-1) compared to 50 microg kg(-1) for microcystin-LR, the data indicate that the non-covalent inhibition of protein phosphatases accounts for most of the harmful effects of microcystins in vivo. A 3-amino-6-hydroxy-2-piperidone containing cyclic peptide, nostocyclin, also isolated from Nostoc sp., was non-toxic and exhibited more than 500-fold less inhibitory potency towards PP1, PP2A, PPP4 and PPP5, consistent with the conclusion that potent inhibition of one or more these protein phosphatases underlies the toxicity of microcystins, both lacking and containing Dhb.

Aminobutyrates↗

Effects of microcystins on the growth and the activity of superoxide dismutase and peroxidase of rape (Brassica napus L.) and rice (Oryza sativa L.).

Microcystins are naturally occurring hepatotoxic cyclic heptapeptides produced by some toxic freshwater cyanobacterial species. In this study, crude extract of toxic cyanobacterial blooms from Dianchi Lake in southwestern China was used to determine the effects of microcystins on rape (Brassica napus L.) and rice (Oryza sativa L.). Experiments were carried out on a range of doses of the extract (equivalent to 0, 0.024, 0.12, 0.6 and 3 microg MC-LR/ml). Investigations showed that exposure to microcystins inhibited the growth and development of both rice and rape seedlings, however, microcystins had more powerful inhibition effect on rape than rice in germination percentage of seeds and seedling height. Microcystins significantly inhibited the elongation of primary roots of rape and rice seedlings. Determination of the activities of peroxidase and superoxide dismutase demonstrated that microcystin stress was manifested as an oxidative stress. Using ELISA, microcystins were examined from the extract of exposed rape and rice seedlings, indicating that consumption of edible plants exposed to microcystins via irrigation route may have health risks. Significantly different levels of recovered microcystins between exposed rice and rape seedlings suggested that there might be different tolerant mechanisms toward microcystins.

Bacterial Toxins↗

Method for detecting classes of microcystins by combination of protein phosphatase inhibition assay and ELISA: comparison with LC-MS.

Depending on the class of microcystin the protein phosphatase inhibition assay shows different sensitivities to different classes of toxin. We have determined that the IC50 values obtained from dose-response curves for the inhibition of the enzyme by micro-cystin LR, nodularin, YR, and RR were 2.2, 1.8, 9 and 175 nM, respectively. When equimolar amounts of these toxins were determined by the ELISA assay with microcystin LR as the standard, the assay showed equivalence in toxin responses. However, when the toxins were determined by the protein phosphatase inhibition assay using microcystin LR as the standard, the ratios of the values determined by PP-2A to ELISA decreased in the order: nodularin (2.23) microcystin LR (1.1)> microcystin YR (0.63)> microcystin RR (0.06). When the ratios for each standard were plotted against the IC50 values, the log-log plot was negative linear, and the lowest value for the IC50 corresponded with the lowest ratio. The differential sensitivity of the PP-2A assay to the various standards was used to establish an indicative toxicity ranking (ITR) where a ranking of 1 (the highest) was assigned to ratios of > or = 0.8 or greater, and 3 (the lowest) to values < or = 0.2. The three ranking classes corresponded to toxin equivalence represented by the four standards. The new method allows not only the determination of microcystin toxins in terms of stoichiometry (ELISA) but also in terms of indicative toxicity. The method can be performed using the same instrument (e.g. multiwell fluorimeter with absorbance capability) and offers an advantage to methods presently used to determine microcystins (e.g. ELISA or LC-MS). The former has the propensity to overestimate toxicity because it measures equivalence to microcystin LR and is a stoichiometric measurement and the latter has the disadvantage in that relatively few of the microcystins that occur naturally are available as standards. The new method was applied to the analysis of sample from lakes and streams from temperate locations and to extracts of cyanobacterial mats from ponds and streams in cold temperature locations.

Bacterial Toxins↗

Responses of antioxidant system in Arabidopsis thaliana suspension cells to the toxicity of microcystin-RR.

Microcystins are cyclic heptapeptide hepatoxins produced by many species of cyanobacteria. The toxic effects and mechanism of microcystins on animals have been well studied both in vivo and in vitro. It was also reported that microcystins had adverse effects on plants. However, to our knowledge, there is no information about the toxic effects and mechanism of microcystins on plant suspension cells. In this study, Arabidopsis thaliana suspension cells were exposed to a range dose of microcystin-RR. Lipid peroxidation, a main manifestation of oxidative damage, was studied and a time- and dose-dependent increase in malondiadehyde was observed. In contrast, glutathione (GSH) levels in the cells decreased after 48 h treatment with 1 and 5mg/L of microcystin-RR. The activities of superoxide dismutase (SOD) and catalase (CAT) increased significantly after 48 h exposure to 1 and 5mg/L of microcystin-RR, but glutathione S-transferase (GST) activity showed no difference compared with the control. These results clearly indicate that microcystin-RR is able to cause oxidative damage in A. thaliana suspension cells. Decrease of GSH content and increases of SOD and CAT activities reveal that the antioxidant system may play an important role in eliminating or alleviating the toxicity of microcystin-RR. The possible toxicity mechanism of microcystin-RR on the A. thaliana suspension cells is also discussed in this paper.

Analysis of Variance↗

Biodegradation of the cyanobacterial toxin microcystin LR in natural water and biologically active slow sand filters.

A bacterium (MJ-PV) previously demonstrated to degrade the cyanobacterial toxin microcystin LR, was investigated for bioremediation applications in natural water microcosms and biologically active slow sand filters. Enhanced degradation of microcystin LR was observed with inoculated (1 x 10(6) cell/mL) treatments of river water dosed with microcystin LR (>80% degradation within 2 days) compared to uninoculated controls. Inoculation of MJ-PV at lower concentrations (1 x 10(2)-1 x 10(5) cells/mL) also demonstrated enhanced microcystin LR degradation over control treatments. Polymerase chain reactions (PCR) specifically targeting amplification of 16S rDNA of MJ-PV and the gene responsible for initial degradation of microcystin LR (mlrA) were successfully applied to monitor the presence of the bacterium in experimental trials. No amplified products indicative of an endemic MJ-PV population were observed in uninoculated treatments indicating other bacterial strains were active in degradation of microcystin LR. Pilot scale biologically active slow sand filters demonstrated degradation of microcystin LR irrespective of MJ-PV bacterial inoculation. PCR analysis detected the MJ-PV population at all locations within the sand filters where microcystin degradation was measured. Despite not observing enhanced degradation of microcystin LR in inoculated columns compared to uninoculated column, these studies demonstrate the effectiveness of a low-technology water treatment system like biologically active slow sand filters for removal of microcystins from reticulated water supplies.

Biodegradation, Environmental↗