Dynamics of microcystins in the mussel Mytilus galloprovincialis.

The accumulation and depuration of hepatotoxins produced by the freshwater cyanobacterium Microcystis aeruginosa in the mussel Mytilus galloprovincialis was studied. Mussels were fed daily 10(5) cells/ml of the toxic cyanobacterium that produces microcystin-LR (MCYST-LR), for four days. After that period animals were placed in toxin free water and were fed the diatom Nitzschia sp. During two weeks the concentration of the toxin in the mussels, as also in their feces and in the water where animals were placed individually during 24 h, were monitored using an ELISA assay. No mussel mortality was registered during the whole experiment. Mussels showed a maximum detectable level of MCYST of 10.7 microg/g mussels dry weight (DW) during the accumulation period, rising to 16.0 microg MCYST/g mussel DW by day two of the depuration period. Then there was a decrease trend with peaks of toxin at days 6, 8, 11 and 14. The rise of the toxin level on day two of the depuration period seems to have been due to the reingestion of contaminated feces. In fact, feces showed high amounts of MCYST during the first days of depuration with a maximum of 140 microg/d DW on day 3. This coincided with a 50% decrease on the detectable toxin in the mussels reflecting the emptiness of their digestive tract. In the water the highest level of the toxin was 2.5 microg MCYST/liter and some toxin peaks were also observed during the depuration period. This fluctuation of the toxin levels in the mussels, feces and water may be related to the renewal of protein phosphatases and subsequent release of unbound toxins. Results show that depuration of MCYST by mussels is not a very rapid process and contamination by feces containing MCYST is likely to occur and increase the persistence of these toxins in the mussels after the bloom disappearance. Monitoring programs for harmful algal blooms usually include only toxic dinoflagellates and diatoms and their toxins in bivalves. Taken into account the present work they should also include hepatotoxins from cyanobacteria, namely in brackish waters such as estuaries of eutrophic rivers in order to avoid human health hazard.     

Impact of a toxic and a non-toxic strain of Microcystis aeruginosa on the crayfish Procambarus clarkii.

The occurrence of cyanobacteria in many water bodies where crayfish such as Procambarus clarkii are abundant leads to the possibility of toxin accumulation and food chain transfer. This paper describes the accumulation and depuration of microcystins from a microcystin and a non-microcystin producing strain of Microcystis aeruginosa, on the survivorship, growth and nutritional status of P. clarkii. Crayfish larvae were resistant to cyanobacteria and their toxins, surviving cyanobacteria densities during acute exposures. Juvenile crayfish tolerated toxic cyanobacteria better than non-toxic ones. This effect was also observed when analysing nutritional status of crayfish fed toxic and non-toxic cyanobacteria with the former having better lipid and protein contents than those fed non-toxic Microcystis. P. clarkii accumulated up to 2.9 microg MCYST/dry crayfish weight and the depuration pattern was similar to that observed for mussels by other authors. Due to the fact that the major part of the toxin is accumulated in the intestine and in the hepatopancreas, there is no significant risk in terms of human health if these parts are removed prior to crayfish consumption. Nevertheless, their use in dairy food and the possible transference of toxins along food chains should not be disregarded.     

Microcystin concentrations and genetic diversity of Microcystis in the lower Great Lakes

The resurgence of Microcystis blooms in the lower Great Lakes region is of great concern to public and ecosystem health due to the potential for these colonial cyanobacteria to produce hepatotoxic microcystins. A survey of Microcystis cell densities and microcystin concentrations during August 2004 showed particularly high concentrations of both cells and toxin in the nearshore regions of Saginaw Bay (Lake Huron) and western Lake Erie, often exceeding the World Health Organization's recommended drinking water limit of 1 microg L(-1). The dominant congener of microcystin in both basins was microcystin-LR (MC-LR), whereas the second most abundant congeners, accounting for up to 20-25% of the total microcystin concentrations, were MC-LA in Saginaw Bay and MC-RR in western Lake Erie. Multiplex PCR assays of Microcystis colonies isolated from these two regions showed that a much greater percentage of the Microcystis colonies from Saginaw Bay carried the mcyB gene necessary for microcystin production, in comparison with those from western Lake Erie. The mcyB genotypes sequenced separated into two distinct phylogenetic clusters, with Microcystis originating from Lake Erie predominantly in one branch and from Saginaw Bay present in both branches. These results indicate that the genetic composition of the bloom could impact the concentrations and congeners of microcystin produced and that the cell count methods currently being used to gauge public health threats posed by Microcystis blooms may not sufficiently assess actual bloom toxicity.     

Experimental model of microcystin accumulation in the liver of Oreochromis niloticus exposed subchronically to a toxic bloom of Microcystis sp

Although accumulation of the liver toxin microcystin in phytoplanktivorous fish has been demonstrated in captive fish and in natural ecosystems, the relation between microcystin in ingested algae and the pattern of buildup of microcystin in fish is poorly known. In this month-long study performed at a Brazilian fish farm, 45 mature Oreochromis niloticus were fed daily with fresh seston periodically dominated by toxic Microcystis sp. Microcystin was measured daily in the food and every 5 days in liver and muscle samples. Control fish received a diet of seston that was low in toxic cyanobacteria. Initially, in treatment ponds, microcystin available for fish increased from 6.5 to 66.9 ng microcystin fish(-1)day(-1), which was accompanied by an increase from 5.5 to 35.4 ng microcysting liver(-1). Microcystin in muscle was below our detection limit of 4 ng g tissue(-1) for the entire study. In the bloom phase, available microcystin reached its highest concentration (4450 ng MC fish(-1)day(-1)) then decreased to 910 ng microcystin fish(-1)day(-1) on day 31. During this period, microcystin reached its highest concentration of 81.6 ng MC g liver(-1) and stayed high until the end of the experiment. A model based on rapid uptake, saturation, and exponential loss was built with these experimental results, and verified with data from the literature. Our model showed that accumulation was up to 50% of ingestion at low doses, but at intermediate doses, the onset of elimination led to a decline of liver burden. Although the accumulation rate confirms the high contamination potential of microcystin, it was balanced by a high depuration rate and this efficient systemic elimination may explain the tolerance of these fish to toxic blooms in the wild.     

Accumulation and depuration of cyanobacterial paralytic shellfish toxins by the freshwater mussel Anodonta cygnea

The increasing frequency by which the production of paralytic shellfish toxins (PST) by freshwater bloom-forming cyanobacteria is being noticed world-wide raises the possibility of PST bioaccumulation by freshwater mussels. This study evaluates PST accumulation and depuration by the freshwater mussel Anodonta cygnea exposed over a 14-day period to high densities (mean = 1.4 x 10(9) cells1(-1), S.D. = 0.29 x 10(9) cellsl(-1)) of the toxic cyanobacterium Aphanizomenon issatschenkoi (corresponding to a mean toxin concentration of 25.5 nmol PSTl(-1), S.D. = 9.9 nmol PSTl(-1)). Mussels were subsequently detoxified either by starvation or by feeding on the non-toxic green-algae Ankistodesmus falcatus. Filter feeding activity and toxin uptake by the mussels were followed by cell counting and toxin analysis in water samples taken before and after each daily water renewal. The accumulation and depuration of PST as well as the anatomical distribution of toxins were monitored throughout the experiment by HPLC analysis of mussel extracts. Mussels fed the toxic cyanobacterium removed on average 65.3% of cells and 40.36% of total PST daily provided. Daily rates of cell clearance (% of initial) were negatively correlated with the amounts of PST daily provided (but not with the amount of cells). This suggests a negative effect of toxins on the feeding behaviour of mussels. Small amounts of toxins could be detected in the mussels after the second day of exposure, reaching a maximum of 26 microg PST100 g(-1) by day 7. The viscera contained the greatest proportion of toxins (78%) at the start of the toxification. However, increasing amounts of PST were found in the remaining tissues (gills, mantle and foot) over time. Toxins detected in the mussel extracts were the same provided in the dietary A. issatschenkoi. Nevertheless, mussels showed a higher proportion of saxitoxin and decarbomoylsaxitoxin and a lower proportion of gonyautoxin-5 than the fed cyanobacterium. Similar depuration efficiencies were observed among starved individuals (6.9% day(-1)) and those fed with A. falcatus (8.2% day(-1)) indicating that both treatments had comparable effects on toxin metabolism. Mussels showed a typical S shaped depuration kinetics curve consisting of a first short period of slow toxin decay followed by a rapid loss and a subsequent slower release of toxins. Trace to undetectable levels of PST were found in mussels after the 14-day depurating period. Although freshwater mussels are not widely consumed by humans, their capacity to accumulate PST points to the risk of PST propagation through the food chain of freshwater ecosystems via filter-feeding mussels.     

Occurrence of toxic blue-green algae in the Kucukcekmece lagoon (Istanbul, Turkey)

The concentration of microcystin (MC) in the Kucukcekmece Lagoon, Istanbul, Turkey, and the physicochemical and biological parameters of water quality were investigated from October 2000 to June 2003. Water samples were collected from surface waters at three sites. Most bloom samples were dominated by Microcystis aeruginosa. The major microcystin variants detected by HPLC-PDA were microcystin-YR and microcystin-LR. Microcystin concentration increased dramatically from early summer to early autumn and thereafter tended to decrease. The toxin concentration found in the filtered samples from surface waters varied between 0.06 and 24.2 microg L(-1) microcystin-LR equivalents. Each year extensive fish mortality was recorded between mid-June and early October, coinciding with heavy algal blooms. A comparison of the conditions associated with cyanotoxin episodes in 2000, 2001, and 2002 showed that the microcystin increase was related to temperature, high concentration of dissolved nutrients, high light intensity (PAR). The highest MC concentrations were recorded at temperatures between 24 degrees C and 28.5 degrees C. Field data showed that the highest MC concentration (>3 microg L(-1)) and the highest cyanobacterial biomass (>30 mg L(-1)) corresponded to a total nitrogen:total phosphorus ratio greater than 7:1. The highest concentrations of M. aeruginosa biomass (173 mg L(-1)) and MC (24.2 microg L(-1) MC-LR equiv.) and the highest salinity (8.8%) were measured concurrently in the lagoon. To our knowledge, this is the first evidence of cyanobacterial toxins in the Kucukcekmece Lagoon.     

Accumulation of microcystins in various organs of the freshwater snail Sinotaia histrica and three fishes in a temperate lake, the eutrophic Lake Suwa, Japan.

So far, there has been only one study to examine microcystin (MC) contents in various organs of snails in a subtropical Chinese lake. In this study, tissue distribution and seasonal dynamics of MC-RR and -LR were investigated in various organs of a freshwater snail (Sinotaia histrica) in a temperate eutrophic lake, Lake Suwa, Japan. Accumulation of microcystins in some fish was also investigated. There was marked temporal variation in the MC content of various organs of the snail. The digestive tract had the highest MC content (mean 9.03 microg g(-1) DW and range 3.74-23.2 microg g(-1) DW), followed by the gonad (mean 6.90 microg g(-1) DW and range 0.07-22.7 microg g(-1) DW) and hepatopancreas (mean 5.38 microg g(-1) DW and range 1.08-8.79 microg g(-1) DW), whereas the foot had the least (mean 2.48 microg g(-1) DW and range 0.04-4.45 microg g(-1) DW). The disappearance of MC-LR in the hepatopancreas indicated that S. histrica is able to depurate MC-LR efficiently. MC-RR was detected in the muscle of three species of fish, with the highest content in Carassius auratus (79.4 microg kg(-1) BW). Because of substantial MC accumulation in these edible aquatic animals in Lake Suwa, it is recommended that regular monitoring of MCs should be undertaken in both cyanobacteria and aquatic animals.     

Organ distribution and bioaccumulation of microcystins in freshwater fish at different trophic levels from the eutrophic Lake Chaohu, China

This article reports the organ distribution and bioaccumulation of hepatotoxic microcystins (MCs) in freshwater fishes at different trophic levels from the large, shallow, eutrophic Lake Chaohu in September 2003, when there were heavy surface blooms of toxic cyanobacteria. Among all fish, intestines and blood had the highest average content of MC-RR + MC-LR (22.0 and 14.5 microg g(-1) DW, respectively), followed by liver, bile, and kidney (7.77, 6.32, and 5.81 microg g(-1) DW, respectively), whereas muscle had the least (1.81 microg g(-1) DW). MC content in muscle was highest in carnivorous fish (Culter ilishaeformis, 2.22 microg g(-1) DW) and omnivorous fish (Carassius auratus, 1.96 microg g(-1) DW) and was lowest in phytoplanktivorous fish (Hypophthalmichthys molitrix, 1.65 microg g(-1) DW) and herbivorous fish (Parabramis pekinensis 0.660 microg g(-1) DW). However, the amount of MC in the gut of H. molitrix (137 microg g(-1) DW) was more than 20 times that in the other fish (<6.50 microg g(-1) DW). The MCs showed a tendency to accumulate up the food chain, and piscivorous fish at the top of the food chain were at high risk of exposure to MCs in Lake Chaohu. Our study is the first to report MC concentrations in the bile and blood of wild fish. One hundred grams of fish muscle would contain 2.64-49.7 microg of MC-LR equivalent, or about 1.3-25 times the recommended tolerable daily intake of MC-LR by humans, indicating that fish are already severely contaminated by MCs and that the local authorities should warn the public of the risk of poisoning by eating the contaminated fish.     

Microcystin-LR degradation utilizing a novel effective indigenous bacterial community YFMCD1 from Lake Taihu

Microcystins (MC) produced by species of cyanobacteria including Microcystis, Anabaena, and Aphanizomenon are a group of monocyclic hepatotoxins posing serious threat to public health. Microcystin-LR (MC-LR) is the most toxic and frequently encountered microcystin variant in the environment, and thus removal of this toxin using bacteria was shown to be a reliable, efficient, and cost-effective method that avoids utilization of chemicals that may produce potentially harmful by-products. The aim of this study was to determine whether a novel indigenous bacterial community designated YFMCD1 was effective in destroying MC. In addition, the influence of environmental factors such as temperature, MC concentration, and pH was examined on the effectiveness of YFMCD1 to degrade MC-LR. MC-degradation products were identified by high performance liquid chromatography coupled with an ultra-high resolution LTQ Orbitrap Velos Pro ETD mass spectrometry equipped with electrospray ionization interface (HPLC-ESI-MS). MC-LR underwent maximal degradation at rate of 0.5 µg/ml/hr with YFMCD1 containing Klebsiella sp. termed YFMCD1-1 or Stenotrophomonas sp. termed YFMCD1-2. Moreover, Adda (3-amino-9-methoxy-2, 6, 8-trimethyl-10-phenyldeca-4, 6-dienoic acid) is a constituent within the MC-LR molecule found to be responsible for biological activity expression and critical for MC-induced toxicity, which is also degraded by YFMCD1. The results showed that YFMCD1 effectively degraded MC-LR. The degradation rate was significantly affected by temperature, pH, and MC-LR concentrations. Data indicate that this bacterial community may prove beneficial in bioremediation of lakes containing MC.     

Short-term uptake of microcystin-LR by Coregonus lavaretus: GST activity and genotoxicity

In the present study, juvenile whitefish weighing 2?g were exposed by force-feeding to two ecologically relevant doses (0.05 and 0.5?μg per fish) of microcystin-LR (MC-LR). Then over 96?h the MC uptake in fish liver and muscle was measured, as the activity of the detoxification enzyme glutathione S-transferase (GST) in the liver, and the genotoxicity impact on red blood cells. Results show that (1) the MC-LR equivalent concentrations increased for both doses and in both organs of whitefish with approximately threefold lower concentrations for the low dose compared to the high dose in both organs and threefold lower concentrations in the muscle compared to the liver for each dose (2) the liver GST activity increased during the first 48?h of exposure with fivefold higher GST activity for the highest dose at 48?h compared to control and (3) MC-LR leads to deoxyribonucleic acid strand breaks that were detected by the comet assay and shown to be partially repaired. This work demonstrates that European whitefish could be impacted by cyanobacteria toxins due to rapid microcystin uptake, especially in the context of chronic contamination, which can occur during long bloom episodes.     

Characterization of microcystin production in an Antarctic cyanobacterial mat community.

Cyanobacteria are well known for their production of non-ribosomal cyclic peptide toxins, including microcystin, in temperate and tropical regions, however, the production of these compounds in extremely cold environments is still largely unexplored. Therefore, we investigated the production of protein phosphatase inhibiting microcystins by Antarctic cyanobacteria. We have identified microcystin-LR and for the first time [D-Asp3] microcystin-LR by mass spectrometric analysis in Antarctic cyanobacteria. The microcystins were extracted from a benthic microbial community that was sampled from a meltwater pond (Fresh Pond, McMurdo Ice Shelf, Antarctica). The extracted cyanobacterial cyclic peptides were equivalent to 11.4 ng MC-LR per mg dry weight by semi-quantitative analyses using HPLC-DAD and the protein phosphatase inhibition assay. Furthermore, we were able to identify the presence of cyanobacterial non-ribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) genes in total DNA extracts from the mat community.     

Initial studies on the occurrence of cyanobacteria and microcystins in Irish lakes

We report the results of a synoptic survey at 14 sites across the north of Ireland undertaken to determine the occurrence of cyanobacteria and their constituent microcystin cyanotoxins. Seven microcystin toxins were tested for, and five of which were found, with MC‐LR, MC‐RR, and MC‐YR being the most prevalent. Gomphosphaeria spp and Microcystis aeruginosa were the most dominant cyanobacterial species encountered. Together with Aphanizomenon flos‐aquae, these were the cyanobacteria associated with the highest microcystin concentrations. The occurrence of several microcystin toxins indicates that there may potentially be more than one cyanobacteria species producing microcystins at many sites. Total microcystin concentrations varied over three orders of magnitude dividing the sites into two groups of high (>1000 ngMC/μgChla, six sites) or low toxicity (<200 ngMC/μgChla, eight sites). © 2010 Wiley Periodicals, Inc. Environ Toxicol, 2010.     

Microcystin-LR causes cytotoxicity effects in rat testicular Sertoli cells

Microcystins (MCs) are produced by cyanobacteria. The most toxic and widely distributed MC is microcystin-LR (MC-LR). The aim of this study was to investigate whether exposure to MC-LR could induce oxidative stress, leading the further toxicity effects on Sertoli cells in vitro. Sertoli cells obtained from rats were cultured with a medium containing 0, 0.5, 5, 50 or 500 nM/l MC-LR. We examined the decrease of mitochondrial membrane potential (MMP), the increase of reactive oxygen species (ROS) production, the increase of lipid peroxidation and decrease of superoxide dismutase (SOD) activity in Sertoli cells after treatment with MC-LR in vitro, and higher expression of caspase-9 and caspase-3, the increase of apoptosis rate. Therefore, we deduced that direct exposure to microcystin-LR could induce oxidative stress generation in Sertoli cells, and subsequently depressed cellular viability and caused cells to undergo apoptosis, resulting in the reproductive toxicity in male rats.     

Microcystins associated with Microcystis dominated blooms in the Southwest wetlands, Western Australia

Potentially toxic cyanobacterial blooms are becoming common in the freshwater wetlands on the Swan Coastal Plain, Western Australia. During summer the dominant bloom-causing species belong to the genera Microcystis and Anabaena and to a lesser extent Aphanizomenon and Nodularia. Although toxic cyanobacteria have been recorded in the Swan-Canning and Peel-Harvey estuaries in Western Australia, very little is known about the blooms in the surrounding freshwater lakes. In this study, a total of 32 natural bloom samples representing 13 lakes were analyzed by HPLC for microcystin (MC)-LR, -RR, and -YR. Twenty-eight samples proved to be toxic. The highest total microcystin concentration ranged from 1645 to 8428.6 microg L(-1), and the lowest concentrations were less than 10 microg L(-1) with some below the detection limit (< 0.05 microg L(-1)). MC-LR (100%) was the predominant microcystin, followed by MC-YR (71.4%) and MC-RR (60.7%). The presence of a Nodularia spumigena bloom in the freshwater Lake Yangebup was associated with the detection of nodularins (1664 microg L(-1)). This is the first study to demonstrate the presence of microcystins and nodularins in urban lakes on the Swan Coastal Plain, Western Australia.     

Toxicology of a Microcystis ichthyoblabe waterbloom from Lake Oued Mellah (Morocco)

In the Lake Oued Mellah cyanobacteria waterblooms occur periodically in late spring and summer with Microcystis ichthyoblabe as the main bloom-forming species. In 1999, a heavy waterbloom of M. ichthyoblabe occurred during May June with a maximal biomass of 298 mg/l. During this period, several bloom samples were collected. The toxicity assessment was done by mouse and brine shrimp (Artemia) bioassays. Apart from the sample collected on 15/06/1999, all the other samples were toxic by mouse bioassay. The LD50 determined by intraperitoneal injection to mice during active cyanobacterial growth and decline phases were 518 and 1924 mgDW/kg respectively. Using Artemia bioassay, the 24hLC50 varied from 6.0 to 40.6 mg/ml and the 48hLC50 ranged from 2.8 to 18.2 mg/ml. The separation and identification of microcystin variants was performed by high performance liquid chromatography-photodiode array detection. Eleven toxins were separated and preliminarily identified as microcystin variants as they exhibit a typical UV spectra like the microcystin-LR standard. The quantification of total microcystins determined by enzyme-linked immunosorbent assay showed that the contents were varied between 0.1 and 0.76 microgram/g DW.     

An investigation of the role of vitamin E in the protection of mice against microcystin toxicity

The presence of cyanobacterial toxins in drinking and recreational waters represents a potential public health risk. Microcystin-LR (MC-LR) is a potent cyclic heptapeptide hepatotoxin produced by the blue-green alga Microcystis aeruginosa. Chemoprotectant studies have indicated that membrane-active antioxidants such as vitamin E may offer protection against microcystin toxicity. This study investigated the effect of vitamin E supplementation on microcystin toxicity in mouse liver. Groups of mice were fed vitamin E supplements (8.33 or 33.3 U/mouse/day) for 4 weeks, with intraperitoneal doses of MC-LR extract (70% LD(50)) every 3 days from day 8. The potential benefits of vitamin E were evaluated based on lipid peroxidation, alanine transaminase (ALT), and glutathione S-transferase (GST) levels. Vitamin E supplementation at 33.3 U/mouse/day offered some protection against lipid peroxidation induced by repeated exposure to MC-LR extract and limited both the toxin-induced increase in ALT leakage and decrease in GST activity. Vitamin E supplementation at 66.6 U/mouse/day significantly increased the time to death and reduced the increase in liver percentage body weight induced in mice given a lethal dose challenge of MC-LR extract. Therefore, vitamin E, taken as a dietary supplement, may have a protective effect against chronic exposure to MC-LR.     

Changes in concentrations of microcystins in rainbow trout, freshwater mussels, and cyanobacteria in Lakes Rotoiti and Rotoehu

Microcystin concentrations in cyanobacteria and their accumulation in rainbow trout (Oncorhynchus mykiss) and freshwater mussels (Hyridella menziesi) in Lakes Rotoiti and Rotoehu (New Zealand) were investigated. Hatchery rainbow trout were added to an enclosure in Lake Rotoiti where concentrations of microcystins in the phytoplankton and cyanobacterial cell concentrations could be closely monitored. Rainbow trout that were free to roam in the entire area of each lake were also included in the study. Freshwater mussels were suspended subsurface in cages in the enclosure. Phytoplankton samples, rainbow trout liver and muscle tissue, and the tissues of mussels were analyzed for microcystins using the ADDA-ELISA method, and selected samples were analyzed using LC-MS. A maximum concentration of microcystins in the phytoplankton samples of 760 microg L(-1) was recorded in Te Weta Bay, Lake Rotoiti, in March 2004. ELISA results confirmed microcystin immunoreactivity in rainbow trout liver and muscle tissues and in freshwater mussels. The microcystin congeners LR, YR, RR, AR, FR, LA, and WR were detected by LC-MS in caged freshwater mussels in Lake Rotoiti but were not detected in either muscle or liver tissue of rainbow trout. The daily tolerable intake limit of microcystins for human consumption recommended by the World Health Organisation is 0.04 microg kg(-1) day(-1). Modeling was carried out for the human intake of microcystin compounds from rainbow trout muscle tissue, and the potential health risks were estimated, assuming the ADDA-ELISA was determining compounds of toxicity equivalent to microcystin-LR.     

Potential synergistic effects of microcystins and bacterial lipopolysaccharides on life history traits of Daphnia galeata raised on low and high food levels

Metastudies have found no consistent effects of the cyanobacterial toxin microcystin on Daphnia, and there are discrepancies between field observations and experiments. Confounding factors include absence or presence of alternative high quality food or the presence of bioactive compounds, other than microcystins in cyanobacteria. Of specific interest are lipopolysaccharides (LPS) on the outer cell wall. LPS may have a number of biological effects, including reduced detoxication of microcystins in plants and animals. When grazing seston in the field, filterfeeders take up heterotrophic bacteria attached to cyanobacteria, as well as free-living bacteria. The LPS produced by heterotrophic bacteria have been shown to be much more harmful than cyanobacterial LPS. We performed two experiments in which we tested for potential synergistic effects between bacterial LPS and microcystins. Full-factorial experiments separated the main effects and interactions between (i) food quantity as well as food quality (addition of the green alga Scenedesmus), (ii) presence or absence of strains that vary in amount and composition of microcystins (microcystin free strain NIVA-CYA43, moderate microcystin producing strain NIVA-CYA140 and high microcystin producing strain PCC7820), and (iii) presence or absence of bacterial LPS on different life history traits of Daphnia galeata. We measured juvenile growth rate, age and size at first reproduction, death before first reproduction and standard carbon content of Daphnia. From the experiments we conclude that microcystin-producing Microcystis had deleterious effects on the life history of D. galeata, but especially when the availability of high quality green algal food was limited in comparison to the supply of microcystin producing strain PCC7820. In the experiment in which PCC7820 was used as microcystin-producing strain, addition of LPS lowered SCC of Daphnia, but had no effects on other life history parameters. The interaction between Microcystis strain, Microcystis concentration and LPS was highly significant in case of PCC7820, but not in case of CYA-140, indicating that the effects of LPS and its interactions with microcystin on Daphnia life history were strongly context dependent.     

Phytotoxic effects of cyanobacteria extract on the aquatic plant Lemna gibba: microcystin accumulation, detoxication and oxidative stress induction

The occurrence of toxic cyanobacteria in the aquatic environment constitutes a serious risk for the ecological balance and the functioning of ecosystems. The presence of cyanotoxins in ecosystems could have eventual adverse effects on aquatic plants, which play an important biological role as primary producers. The original aim of this study was to investigate microcystin (MC) accumulation, detoxication and oxidative stress induction in the free-floating aquatic vascular plant Lemna gibba (Duckweed, Lemnaceae). Experiments were carried out with a range of MC levels, obtained from toxic Microcystis culture extracts (0.075, 0.15, 0.22 and 0.3 microg equiv.MC-LR mL(-1)). During chronic exposure of the plant to MC, we examined the growth, photosynthetic pigment contents and also the physiological behavior related to toxin accumulation, possible biodegradation and stress oxidative processes of L. gibba. For the last reason, changes in peroxidase activity and phenol compound content were determined. This is a first report using phenol compounds as indicators of biotic stress induced by MC contamination in aquatic plants. Following MC exposure, a significant decrease of plant growth and chlorophyll content was observed. Also, it was demonstrated that L. gibba could take up and bio-transform microcystins. A suspected MC degradation metabolite was detected in treated Lemna cells. In response to chronic contamination with MCs, changes in the peroxidase activity and qualitative and quantitative changes in phenolic compounds were observed after 24h of plant exposure. The physiological effects induced by chronic exposure to microcystins confirm that in aquatic ecosystems plants coexisting with toxic cyanobacterial blooms may suffer an important negative ecological impact. This may represent a sanitary risk due to toxin bioaccumulation and biotransfer through the food chain.     

Seasonal dynamics of the hepatotoxic microcystins in various organs of four freshwater bivalves from the large eutrophic lake Taihu of subtropical China and the risk to human consumption

So far, little is known on the distribution of hepatotoxic microcystin (MC) in various organs of bivalves, and there is no study on MC accumulation in bivalves from Chinese waters. Distribution pattern and seasonal dynamics of MC-LR, -YR and -RR in various organs (hepatopancreas, intestine, visceral mass, gill, foot, and rest) of four edible freshwater mussels (Anodonta woodiana, Hyriopsis cumingii, Cristaria plicata, and Lamprotula leai) were studied monthly during Oct. 2003-Sep. 2004 in Lake Taihu with toxic cyanobacterial blooms in the summer. Qualitative and quantitative determinations of MCs in the organs were done by LC-MS and HPLC. The major toxins were present in the hepatopancreas (45.5-55.4%), followed by visceral mass with substantial amount of gonad (27.6-35.5%), whereas gill and foot were the least (1.8-5.1%). The maximum MC contents in the hepatopancreas, intestine, visceral mass, gill, foot, and rest were 38.48, 20.65, 1.70, 0.64, 0.58, and 0.61 microg/g DW, respectively. There were rather good positive correlation in MC contents between intestines and hepatopancreas of the four bivalves (r=0.75-0.97, p<0.05). There appeared to be positive correlations between the maximum MC content in the hepatopancreas and the delta13C (r=0.919) or delta15N (r=0.878) of the foot, indicating that the different MC content in the hepatopancreas might be due to different food ingestion. A glutathione (GSH) conjugate of MC-LR was also detected in the foot sample of C. plicata. Among the foot samples analyzed, 54% were above the provisional WHO tolerable daily intake (TDI) level, and the mean daily intakes from the four bivalves were 8-23.5 times the TDI value when the bivalves are eaten as a whole, suggesting the high risk of consuming bivalves in Lake Taihu.