Μg Microcystin Research Articles

Changes in Extracellular Microcystins (MCs) Accompanying Algae/Cyanobacteria Removal during Three Representative Algae/Cyanobacteria Inactivation Processes and an MC Diffusion Model in Still Water

This study addresses the lack of comparative research on algae/cyanobacteria elimination technologies in the existing literature. Our investigation focused on evaluating the performance of three commonly used algae/cyanobacteria removal processes: ultrasound (20.8 kHz), copper sulfate and biotic algicide (Bacillus subtilis). The evaluation considered both algae/cyanobacteria removal efficacy and the consequent changes in extracellular microcystins (MCs). To achieve this, we employed real eutrophic water as the test water. The ultrasound treatment demonstrated effective algae/cyanobacteria removal, with an average rate of algae/cyanobacteria decreasing (RAD) ranging from 0.50 to 0.99 µg chlorophyll a per liter per minute (µg chlorophyll a/L·min). On the other hand, the copper sulfate and biotic algicide treatments exhibited relatively lower rates of algae/cyanobacteria removal, with average RAD values of 0.21 to 0.38 µg chlorophyll a per liter per day (µg chlorophyll a/L·d) and 0.10 to 0.13 µg chlorophyll a per liter per day (µg chlorophyll a/L·d), respectively. Moreover, we observed significant increases in extracellular MCs in the ultrasound and copper sulfate treatments. The corresponding values of the increment of extracellular MCs accompanying removal per microgramme (µg) chlorophyll a (IEMARMC) were 0.34 to 2.43 µg MCs per µg chlorophyll a (µg MCs/µg chlorophyll a) and 18.13 to 185.08 µg MCs per µg chlorophyll a (µg MCs/µg chlorophyll a), respectively. However, in certain conditions where sufficient dosages (0.5 to 2 mg/L) and reaction time (≥8 days) are provided, the biotic algicide treatment could result in a decrease in MCs compared to an untreated control group (IEMARMC: −43.94 to −32.18 µg MCs per µg chlorophyll a). This suggests that the biotic algicide effectively degraded the MCs. In addition, we developed a one-dimensional MC diffusion model in still water based on Newton’s second law, which exhibits excellent simulation capabilities.

Hepatotoxic shellfish poisoning: Accumulation of microcystins in Eastern oysters (Crassostrea virginica) and Asian clams (Corbicula fluminea) exposed to wild and cultured populations of the harmful cyanobacteria, Microcystis

The Asian clam (Corbicula fluminea) and eastern oyster (Crassostrea virginica) are important resource bivalves found in and downstream of waterways afflicted with cyanobacterial harmful algae blooms (CHABs), respectively. This study examined the potential for C. fluminea and C. virginica to become vectors of the hepatotoxin, microcystin, from the CHAB Microcystis. Laboratory experiments were performed to quantify clearance rates, particle selection, and accumulation of the hepatotoxin, microcystin, using a microcystin-producing Microcystis culture isolated from Lake Erie (strain LE-3) and field experiments were performed with water from Microcystis blooms in Lake Agawam, NY, USA. Clearance rates of Microcystis were faster (p<0.05) than those of Raphidocelis for C. fluminea, while C. virginica cleared Microcystis and Tisochrysis at similar rates. For both bivalves, clearance rates of bloom water were slower than cultures and clams displayed significantly greater electivity for green algae compared to wild populations of cyanobacteria in field experiments while oysters did not. In experiments with cultured Microcystis comprised of single and double cells, both bivalves accumulated >3 µg microcystins g − 1 (wet weight) in 24 – 72 h, several orders of magnitude beyond California guidance value (10 ng g − 1) but accumulated only up to 2 ng microcystins g − 1 when fed bloom water dominated by large Microcystis colonies for four days. For Asian clams, clearance rates and tissue microcystin content decreased when exposed to toxic Microcystis for 3 – 4 days. In contrast, eastern oysters did not depurate microcystin over 3 - 4-day exposures and accumulated an order of magnitude more microcystin than clams. This contrast suggests Asian clams are likely to accumulate minor amounts of microcystin by reducing clearance rates during blooms of Microcystis, selectively feeding on green algae, and depurating microcystin whereas oysters are more likely to accumulate microcystins and thus are more likely to be a vector for hepatotoxic shellfish poisoning in estuaries downstream of Microcystis blooms.

Interannual and Spatial Variability of Cyanotoxins in the Prespa Lake Area, Greece

The Prespa Lakes area in Greece—comprised partly of lake Great and lake Lesser Prespa and the Vromolimni pond—has a global importance for biodiversity. Although the waters show regular cyanobacteria blooms, assessments of water quality threats are limited. Samples collected in 2012 revealed scattered and low microcystin (MC) concentrations in Great Prespa (&lt;0.2 μg MC L−1) whereas considerable spatial heterogeneity in both total chlorophyll (2.4–93 µg L−1) and MC concentrations (0.04–52.4 µg MC L−1) was detected in Lesser Prespa. In 2013, there was far less spatial variability of MC concentrations in Lesser Prespa (0.4–1.53 µg L−1), however in 2014, increased concentrations were detected near the lakeshore (25–861 µg MC L−1). In Vromolimni pond the MC concentrations were on average 26.6 (±6.4) µg MC L−1 in 2012, 2.1 (±0.3) µg MC L−1 in 2013 and 12.7 (±12.5) µg MC L−1 in 2014. In 2013, no anatoxins, saxitoxins, nor cylindrospermopsins were detected in Lesser Prespa and Vromolimni waters. Tissue samples from carps, an otter and Dalmatian Pelicans contained 0.4–1.9 µg MC g−1 dry weight. These results indicate that cyanotoxins could be a threat to the ecosystem functions of particularly Lesser Prespa and Vromolimni.

Trophic Interactions between Anadromous Juvenile Alewife (Alosa pseudoharengus) and Cyanobacterial Populations in a Shallow Mesotrophic Pond

Alosa pseudoharengus is an anadromous fish that migrates from marine to freshwaters to spawn. The early larval and juvenile forms are known to be planktivorous, where heavy feeding upon their preferred food source of large crustacean zooplankton often results in changes to composition and size structure within this trophic guild which in turn can result in shifts within the trophic spectrum and a classic trophic cascade. In this study of Lower Mill Pond, Brewster MA, we evaluated the feeding strategy of juvenile Alosa to determine whether juvenile alewife switches to feeding largely on cyanobacteria and whether cyanotoxins microcystin (MC) and β-methlyamino-L-alanine (BMAA) bioaccumulate in their muscle tissue. Within 15 - 30 days of their estimated spawning date, overexploitation of crustacean zooplankton resulted in a shift from planktivory to benthic detritivory for the majority of their life history, although this did not reduce their condition based on weight-length relationships (Log Wwt. = -5.503 + (3.101 × Log Length). Mean MC (0.003 μg·g-1 dwt) and BMAA (4.49 μg·g-1 dwt) concentrations in the muscle tissue of out-migrating juveniles were presumably derived from benthic subsidies, exporting freshwater cyanotoxins and creating a potential transfer to consumer of 0.0012 μg MC and 1.85 μg BMAA. Biodilution of MC and biomagnification of BMAA were observed. Depletion of the crustacean biomass by >95% resulted in an increase in the rotifer biomass, where Log crustacean (μg·L-1 dwt) = - 5.642 - (7.976 × Log rotifer (μg·L-1 dwt), and an increase in the amount of potentially edible <50 μm cyanobacterial biomass (r(8) = - 0.676, p = 0.046). A secondary cascade appears to have been maintained via invertebrate planktivory by Chaoborus spp.; however for a period of time edible cyanobacteria growth exceeded grazing pressure, resulting in a bloom of edible cyanobacteria. Continued grazing resulted in a shift to larger, inedible cyanobacterial communities where late season (October) surface accumulations were observed. The mass occurrence of juvenile Alosa pseudoharengus appears to be coupled to the sequential increases of cyanobacterial biomass via its influence on the trophic spectrum. Overall, the rotifer biomass (μg·L-1) was positively correlated with MC (pg·mL-1) (r(8) = 0.577, p = 0.104), and negatively correlated with BMAA (μg·L-1) (r(8) = - 0.388, p = 0.373) in the edible cyanobacterial fraction of the water column, although neither of these were significant.

Ecotoxicity assessment of microcystins from freshwater samples using a bioluminescent cyanobacterial bioassay

The hepatotoxic cyanotoxins microcystins (MCs) are emerging contaminants naturally produced by cyanobacteria. Yet their ecological role remains unsolved, previous research suggests that MCs have allelopathic effects on competing photosynthetic microorganisms, even eliciting toxic effects on other freshwater cyanobacteria. In this context, the bioluminescent recombinant cyanobacterium Anabaena sp. PCC7120 CPB4337 (hereinafter Anabaena) was exposed to extracts of MCs. These were obtained from eight natural samples from freshwater reservoirs that contained MCs with a concentration range of 0.04–11.9 μg MCs L−1. MCs extracts included the three most common MCs variants (MC-LR, MC-RR, MC-YR) in different proportions (MC-LR: 100–0%; MC-RR: 100–0%; MC-YR: 14.2–0%). The Anabaena bioassay based on bioluminescence inhibition has been successfully used to test the toxicity of many emerging contaminants (e.g., pharmaceuticals) but never for cyanotoxins prior to this study. Exposure of Anabaena to MCs extracts induced a decrease in its bioluminescence with effective concentration decreasing bioluminescence by 50% ranging from 0.4 to 50.5 μg MC L−1 in the different samples. Bioluminescence responses suggested an interaction between MCs variants which was analyzed via the Additive Index method (AI), indicating an antagonistic effect (AI < 0) of MC-LR and MC-RR present in the samples. Additionally, MC extracts exposure triggered an increase of intracellular free Ca2+ in Anabaena. In short, this study supports the use of the Anabaena bioassay as a sensitive tool to assess the presence of MCs at environmentally relevant concentrations and opens interesting avenues regarding the interactions between MCs variants and the possible implication of Ca2+ in the mode of action of MCs towards cyanobacteria.

Grazing of the copepod Cyclops vicinus on toxic Microcystis aeruginosa: potential for controlling cyanobacterial blooms and transfer of toxins

Abstract Grazing of zooplankton on phytoplankton may contribute to a reduction of harmful cyanobacteria in eutrophic waters. However, the feeding capacity and interaction between zooplankton and toxic cyanobacteria vary among grazer species. In this study, laboratory feeding experiments were designed to measure the grazing rate of the copepod Cyclops vicinus on Microcystis aeruginosa and the potential microcystin (MC) accumulation in the grazer. Copepods were fed a mixed diet of the edible green alga Ankistrodesmus falcatus and toxic M. aeruginosa for 10 days. The results showed that C. vicinus efficiently ingested toxic Microcystis cells with high grazing rates, varying during the feeding period (68.9–606.3 Microcystis cells animal-1 d-1) along with Microcystis cell density. Microcystis cells exhibited a remarkable induction in MC production under grazing conditions with concentrations 1.67–12.5 times higher than those in control cultures. Furthermore, C. vicinus was found to accumulate MCs in its body with concentrations increasing during the experiment (0.05–3.21 μg MC animal-1). Further in situ studies are needed to investigate the ability of Cyclops and other copepods to assimilate and detoxify MCs at environmentally relevant concentrations before deciding on the biocontrol of Microcystis blooms by copepods.

A Systematic Literature Review for Evidence of Aphanizomenon flos-aquae Toxigenicity in Recreational Waters and Toxicity of Dietary Supplements: 2000⁻2017.

Previous studies of recreational waters and blue-green algae supplements (BGAS) demonstrated co-occurrence of Aphanizomenon flos-aquae (AFA) and cyanotoxins, presenting exposure risk. The authors conducted a systematic literature review using a GRADE PRISMA-p 27-item checklist to assess the evidence for toxigenicity of AFA in both fresh waters and BGAS. Studies have shown AFA can produce significant levels of cylindrospermopsin and saxitoxin in fresh waters. Toxicity studies evaluating AFA-based BGAS found some products carried the mcyE gene and tested positive for microcystins at levels ≤ 1 μg microcystin (MC)-LR equivalents/g dry weight. Further analysis discovered BGAS samples had cyanotoxins levels exceeding tolerable daily intake values. There is evidence that Aphanizomenon spp. are toxin producers and AFA has toxigenic genes such as mcyE that could lead to the production of MC under the right environmental conditions. Regardless of this ability, AFA commonly co-occur with known MC producers, which may contaminate BGAS. Toxin production by cyanobacteria is a health concern for both recreational water users and BGAS consumers. Recommendations include: limit harvesting of AFA to months when toxicity is lowest, include AFA in cell counts during visible blooms, and properly identify cyanobacteria species using 16S rRNA methods when toxicity levels are higher than advisory levels.

Germination and seedling growth of Brassica juncea and Medicago sativa under exposure to microcystinscontaminated water from Dau Tieng reservoir

Occurrence of toxic cyanobacteria and their toxins has been a root of many consequently serious problems to aquatic plants, animals, and human in the world. However, the toxicity of cyanobacterial toxins from Vietnam waters to the germination and growth of plants has not studied yet. In this study, we investigated the effects of microcystins (MC) (at the concentrations of 20 and 200 µg MC L–1) from field water and crude extract of cyanobacterial scum (mainly Microcystis spp.) from the Dau Tieng Reservoir, on the germination and seedling of Brassica juncea and Medicago sativa during 7 days. The results showed that MC inhibited the germination of the exposed seeds from 10 – 35% compared to the control. The fresh weight of exposed B. juncea and M. sativa compared to the control was from 44 – 89% and from 55 – 90%, respectively. Root length of the plants reduced from 12 – 73% for B. juncea and from 17 – 54% for M. sativa during toxin exposures. Similarly, the decrease of the shoot length, compared to the control, was from 20 – 80% for B. juncea and 13 – 52% for M. sativa. Therefore, the use of water contaminated with MC for irrigation may negatively influence on the quality and yield of agricultural crops. It could be inferred that there should be an uptake of MC by the exposed seeds and th toxin might be transferred to the next consumers via the food chain.

Phytoplankton Community Shifts and Harmful Algae Presence in a Diversion Influenced Estuary

River water entering estuaries affects the physical and chemical environment at irregular intervals creating a highly dynamic aquatic habitat. Phytoplankton are important primary producers in estuaries that respond quickly to their changing environment. Over a 12-month period, the phytoplankton response was examined in terms of biomass, abundance, community composition, and potential phycotoxin production to seasonal changes in river input entering the Breton Sound Estuary in Louisiana. Chlorophyll a (chl a) measurements estimated phytoplankton biomass and light microscopy identified phytoplankton abundance and community composition. Phycotoxins were measured using ultra-sensitive enzyme-linked immunosorbent assay (ELISA). During the study period, chl a and cell abundances exhibited an inverse relationship with the seasons, which were determined by temperature and diversion input rates. The community was dominated by cyanobacteria for most of the year, mainly corresponding to the warm-low flow season. For the rest of the year, cyanobacteria decreased while chlorophytes and centric diatoms increased to approximately equal contributions. Overall, the phytoplankton community composition was most commonly moderated by temperature, salinity, and dissolved inorganic nitrogen (DIN) availability using BIO-ENV. Microcystins (MCs) were detected throughout the year ranging from below detection to 2.92 μg MCs l−1. MC levels were highest during the warm-low flow season and toward the lower estuary. The detection of MCs for the first time in the Breton Sound Estuary illustrates a potential risk to human health and economically important estuarine food webs.

Accumulation and effects of cyanobacterial microcystins and anatoxin-a on benthic larvae of Chironomus spp. (Diptera: Chironomidae)

Larvae of Chironomidae are distributed world-wide and are very abundant in eutrophic water bodies affected by cyanobacterial blooms. However, there is little information on the effect of cyanobacteria and their metabolites on these aquatic organisms. Our studies revealed that benthic species of Chironomus inhabiting a hypertrophic lake where blooms of microcystin (MC) and/or anatoxin-a (ANTX)-producing filamentous Planktothrix agardhii, Dolichospermum spp. and Cuspidothrix issatschenkoi occur, fed on these cyanobacteria and accumulated cyanotoxins. Up to 3.2 µg MCs g-1 F.W. and up to 185 µg ANTX g-1 F.W. were detected. Of the four MC variants detected in the cyanobacterial biomass [Asp3, Dhb7]MC-RR and MC-LR prevailed, whereas in the larvae it was [Asp3, Dhb7]MC-RR and MC-LA. The effect of pure MC-LR and ANTX as well as crude extracts of MC-producing P. agardhii and ANTX-producing D. lemmermannii on lake and riverine larvae of Chironomus spp. was also compared. The assays revealed that pure cyanotoxins (concentrations: 0.83-3.32 mg L-1) were generally less toxic to riverine larvae than cyanobacterial extracts containing approximately 10-times less toxins. The survival of both the lake and riverine Chironomus larvae did not decrease when exposed to environmentally relevant concentrations of cyanotoxins (< 0.20 mg L-1). The larvae were also highly resistant to higher amounts (up to 0.35 mg ANTX L-1 and 0.42 mg MCs L-1) of extracellular toxins. In the natural environment, Chironomus larvae exposed to toxins contained in cyanobacterial prey, dissolved in water and/or bound to bottom sediments may be very important vectors of cyanotoxins to higher levels in aquatic food chains. To the best of our knowledge, this is the first report on the accumulation of ANTX and effects of cyanotoxins on Chironomus larvae.

Bioacumulation of Cyanotoxins in Hypophthalmichthys molitrix (Silver Carp) in Paranoá Lake, Brasilia-DF, Brazil

Three individuals of silver carp (Hypophthalmichthys molitrix) were collected biweekly from Paranoá Lake (DF, Brazil) for analysis of microcystin (MC) concentrations in their muscle and liver tissue. Analysis by high performance liquid chromatography and mass spectrometry revealed MC masses and fragmentation patterns that were identified as MC-LR (995.04m/z), MC-LA (909.01m/z) and an unknown MC (987.07m/z). Concentrations were calculated as MC-LR equivalents using a calibration curve prepared with a standard of MC-LR. May/06 was the month with the highest MC-LR equivalent concentrations in muscle and liver (3.83±2.78, and 12.94±10.51μgg(-1), respectively). Our results show that during the drought months (April-September), consumption of fish with these MC concentrations would result in exposure to MCs that greatly exceed the World Health Organization's recommended tolerable daily intake limit of 0.04μg MC kg(-1) body weight.

Grazing on toxic cyanobacterial blooms by tadpoles of edible frog Rana grylio

SUMMARYTadpoles of Rana grylio were raised as edible frogs in fishponds of Guanqiao in Wuhan City, Hubei, China, during cyanobacterial blooms from June to October. The dominant cyanobacterial species was Microcystis, which was found to be lethally toxic by intraperitoneal (i.p.) mouse bioassay. Little is known about the effect of tadpoles on toxic cyanobacterial blooms. To evaluate the potential of the tadpoles to graze on cyanobacterial blooms, the tadpoles were fed on Microcystis collected from the field in the laboratory. The Microcystis cells decreased from 1.19 × 107 cells mL−1 to 3.23 × 106 cells mL−1, with a sharp reduction of 73% of the initial Microcystis population observed in the first 24 h after introduction of the tadpoles. The ponds containing tadpoles had a markedly lower density of Microcystis than those lacking tadpoles. Tadpoles exposed to either cultured Microcystis aeruginosa (NIES–90, 2.768 µg microcystins mg–1 dw–1) cells or lysed M. aeruginosa cells grew well, however, indicating that they were unaffected by Microcystis toxins. We found a significant increase in tadpole body weight after feeding on either field Microcystis or cultured M. aeruginosa. The mean increase in individual body weight was 20 mg day−1 when fed on Microcystis from the pond, and 7 mg day−1 when fed on M. aeruginosa from culture. Our study strongly suggested that there is a direct trophic relationship between R. grylio tadpoles and toxic Microcystis blooms and they possess the potential to graze on toxic Microcystis. The results imply that R. grylio tadpoles may play an important ecological role in reducing toxic cyanobacterial blooms caused by Microcystis.

Evaluating the potential risk of microcystins to blue crab ( Callinectes sapidus) fisheries and human health in a eutrophic estuary

Toxin producing cyanobacteria commonly inhabit fresh waters and brackish estuaries, and blooms of these species may be escalating worldwide due to eutrophication. The most common cyanobacterial toxins occurring in fresh and brackish waters are microcystins (MC), which are known to accumulate in aquatic organisms. Prey preference for filter-feeding organisms, such as clams and mussels, make the blue crab, Callinectes sapidus, a candidate for microcystin contamination, therefore making this commercially important edible crab species a potential vector of these toxins to humans. The present study was conducted in a hyper-eutrophic freshwater lake, Lac des Allemands, located in the Barataria estuary system of southeastern Louisiana, and was aimed at documenting the presence and abundance of toxic cyanobacteria and assessing microcystin concentrations in surface water and blue crabs taken from this region. Microcystis sp. were the dominant cyanobacteria, with alternating blooms of Microcystis and Anabaena spp. occurring during the 8-month study. Enzyme-linked immunosorbent assay (ELISA) was used to evaluate concentrations of microcystins from surface water and hepatopancreas, viscera, and muscle tissues of blue crabs. The highest concentration of microcystins found in surface water (1.42 μg MC l −1) was above the tolerable daily intake (TDI) guideline for microcystins in drinking water (1.0 μg MC l −1) set by the World Health Organization (WHO). Highest concentration of microcystins occurring in crab tissue were 820 μg MC kg −1 in hepatopancreas, 65 μg MC kg −1 in viscera, and 105 μg MC kg −1 in muscle, which were close to or exceeding the WHO-TDI guidelines for human consumption (0.04 μg MC kg −1 body weight day −1) based on human body weight and amount of crab tissue consumed. This study documents the presence of microcystins in both surface water and blue crab tissue and therefore, demonstrates the potential for Microcystis and Anabaena blooms to produce toxins that may be accumulated in the tissues of blue crabs and transferred to higher level consumers, including humans.

Analysis of dissolved microcystins in surface water samples from Kovada Lake, Turkey

Dissolved (extracellular) microcystin (MC) concentrations were determined at 3 sampling stations on Lake Kovada, Turkey. The dominant species of cyanobacteria found in August and September of 2006 were Microcystis aeruginosa, Synechococcus sp., Phormidium limosum, Phormidium formosa and Planktothrix limnetica. MC concentrations in water were measured by ELISA and MC variants were examined by HPLC-PDA. Quantitative analysis by HPLC indicated that five MC variants (MC–LR, –RR, –LA, –LW, –LF) were identified in water samples from Kovada Lake. The maximum concentration of dissolved MC–LW was 98.9 µg l − 1 in October. MC–LR was only detected in May at a concentration of 0.5 µg l − 1 . The cross reactivity of the antibody (MC10E7) to variants such as MC–LA MC–LW & MC–LF was low. Hence the results determined by ELISA were lower than those determined by HPLC in September and October samples due to differences in the specificity of the antibody to MC variants. Total extraceullar MCs was quantified by ELISA and ranged from 0.73 to 48.5 µg MC–LR equivalents l − 1 , which in some cases exceeded the WHO provisional Guideline Value for MC–LR in drinking water. This study confirms that the lakes of Turkey should be monitored for toxic cyanobacteria and for MCs to avoid or reduce the potential exposure of people to these health hazards.

Effects of dietary cyanobacteria of two different sources on growth and recovery of hybrid tilapia ( Oreochromis niloticus × O. aureus)

A 115 days feeding trial was conducted to evaluate the effect of dietary cyanobacteria on growth, microcystins (MCs) accumulation in hybrid tilapia ( Oreochromis niloticus × O. aureus) and the recovery when the fish were free of cyanobacteria. Three experimental diets were formulated: the control (cyanobacteria free diet); one test diet with cyanobacteria from Lake Taihu (AMt, 80.0 μg MCs g −1 diet) and one with cyanobacteria from Lake Dianchi (AMd, 410.0 μg MCs g −1 diet). Each diet was fed to fish for 60 days and then all fish were free of cyanobacteria for another 55 days. A significant increase in feeding rate (FR) was observed in fish fed AMd diet after a first 30-day exposure (1 st EP), and in fish fed both AMt diet and AMd diet after a second 30-day exposure (2 nd EP). Specific growth rates (SGR) of fish fed AMt diet and AMd diet were both obviously affected after the first 30-day exposure, but SGR was only significantly affected in fish fed AMt diet after the second 30-day exposure. After a 55-day recovery, there were no significant differences among diets in the indices mentioned above. Much higher concentrations of MCs were accumulated in tissues of all fish exposed to cyanobacteria. After the 55-day recovery, MC concentrations in fish tissues were significantly lower than those on day 60.

Intraperitoneal injection of extracted microcystins results in hypovolemia and hypotension in crucian carp ( Carassius auratus)

Circulatory responses of crucian carp injected intraperitoneally with extracted microcystins (MCs) were studied at sublethal and lethal doses (150 and 600 μg MC kg −1 body mass, respectively). Mean arterial blood pressure (MAP), heart rate, hematocrit (Hct), red blood cell (RBC) counts, and circulating blood volume (BV) were assayed at 0, 1, 3, 12, 24, and 48 h post-toxin administration. MAP decreased significantly in a dose-dependent manner over time. Within the 48-h test period, the lethal dose as well as the sublethal dose resulted in a steady decline of MAP without recovery. Heart rate significantly increased within 24 h post-injection as blood pressure significantly dropped, then showed a terminal decline to the control level. The dose-dependent decreases in BV and Hct were directly related to the drop in MAP. Intraperitoneal injection of a lethal dose of MCs led to hepatic and gill hemorrhage. Consequently, crucian carp given MCs suffered from hypovolemic hypotensive shock.

Variation in peptide net production and growth among strains of the toxic cyanobacterium Planktothrix spp.

Cyanobacteria frequently form mass developments in surface waters. Populations consist of strains that differ in the production of bioactive peptides, e.g. microcystins (MC) inhibiting protein phosphatases 1 and 2A and anabaenopeptins (APN) inhibiting carboxypeptidases. Forty-nine strains (18 green-pigmented (phycocyanin-rich) P. agardhii strains and 31 red-pigmented (phycoerythrin-rich) P. rubescens strains) of the filamentous cyanobacterium Planktothrix (Anagnostidis et Komarék) were analysed for their MC and APN net production rates. These rates were compared with (i) the pigmentation, (ii) the proportion of extra- and intracellular peptide concentrations and (iii) the cellular growth rates under standardized laboratory conditions. Excluding the strains lacking MC and APN, the MC and APN contents varied up to 14-fold and 12-fold, each. The variation in minimum and maximum peptide content (0.32–4.51 µg MC mg−1 dry weight; 0.85–10.32 µg APN mg−1 dry weight) exceeded the variation found for chlorophyll a (4.8–16.9 µg mg−1 dry weight). The extracellular proportions of MC (0–62%) and APN (0–58%) varied among strains, however, on average, proportions of extracellular MC and APN were low (MC: 8.8 ± (1 SE) 1.9%, APN: 8.4 ± 1.8%). Among all strains cellular growth rates showed a 5-fold variation (0.07–0.33 doublings of dry weight.day−1) and were found independent of the pigmentation and the peptide net production rate. It is concluded that the MC and APN net production rate is not causally related to the cell-division cycle and the synthesis of highest amounts of MC and APN does not constrain cell division.

Seasonal occurrence and toxicity of Microcystis spp. and Oscillatoria tenuis in the Lebna Dam, Tunisia

Physicochemical and biological water quality, including the total microcystin concentrations, was investigated for the first time from January to December 2005 in the Lebna Dam, Tunisia. Microcystin levels and characterization of the different microcystin variants present were measured by protein phosphatase (PP2A) inhibition assays and by LC/MS/MS, respectively. Nutrient values were high, with total inorganic nitrogen and phosphorus concentrations ranging from 0.05 to 8.4 mg L −1 and from 0.03 to 1.37 mg L −1, respectively. However, the chlorophyll- a concentrations were very low with a peak (5.32 μg L −1) on 20 September 2005 at 9 m depth water samples. Microscopic examination of the phytoplankton samples showed the dominance in the autumn of three morphospecies of the genus Microcystis and the species Oscillatoria tenuis. The total (particulate and dissolved) microcystin concentrations at the surface and at 9 m depth water samples ranged between 0.008 and 1.73, and 0.005 and 5.57 μg microcystin (MC)-LR equivalent L −1, respectively, with a peak on 20 September. The presence of the microcystin synthetase genes ( mcyA, - B, and - C) in the lysates of the three morphospecies of the genus Microcystis and the species O. tenuis indicated that these species were responsible for the microcystin production in this system. The analysis of the field cyanobacterial sample extract containing these species by LC/MS/MS revealed the presence of two microcystin variants: microcystin-LR (MC-LR) and microcystin-YR (MC-YR).

Effects of microcystins on broccoli and mustard, and analysis of accumulated toxin by liquid chromatography–mass spectrometry

Microcystins (MCs) are cyclic heptapeptides and protein phosphatase inhibitors produced by many species of cyanobacteria. MCs have been shown to cause adverse effects on animals as well as plants and therefore methods are needed for analysing MCs in different matrices. We assessed the effects of MC exposure on broccoli ( Brassica oleracea var. italica) and mustard ( Sinapis alba) by watering the seedlings with water containing 0, 1 or 10 μg MCs L −1 (concentrations typically found in natural waters). Morphological characteristics, chlorophyll concentrations and chlorophyll fluorescence were investigated, but the only distinct difference compared to control plants was a slight (<10%) growth inhibition seen in broccoli. Afterwards the MC concentration of selected plant samples was quantitated using liquid chromatography–mass spectrometry. Among the four MC variants present in the exposure mixture, only MC-LR was clearly detectable, and the toxin was found only in the roots of broccoli and mustard. The detected MC-LR concentrations ranged from 0.9 to 2.6 ng (g fresh weight) −1.

Occurrence and elimination of cyanobacterial toxins in drinking water treatment plants

Toxin-producing cyanobacteria (blue-green algae) are abundant in surface waters used as drinking water resources. The toxicity of one group of these toxins, the microcystins, and their presence in surface waters used for drinking water production has prompted the World Health Organization (WHO) to publish a provisional guideline value of 1.0 μg microcystin (MC)-LR/l drinking water. To verify the efficiency of two different water treatment systems with respect to reduction of cyanobacterial toxins, the concentrations of MC in water samples from surface waters and their associated water treatment plants in Switzerland and Germany were investigated. Toxin concentrations in samples from drinking water treatment plants ranged from below 1.0 μg MC-LR equiv./l to more than 8.0 μg/l in raw water and were distinctly below 1.0 μg/l after treatment. In addition, data to the worldwide occurrence of cyanobacteria in raw and final water of water works and the corresponding guidelines for cyanobacterial toxins in drinking water worldwide are summarized.