Sporadic distribution and distinctive variations of cylindrospermopsin genes in cyanobacterial strains and environmental samples from Chinese freshwater bodies.

Increasing reports of cylindrospermopsins (CYNs) in freshwater ecosystems have promoted the demand for identifying all of the potential CYN-producing cyanobacterial species. The present study explored the phylogenetic distribution and evolution of cyr genes in cyanobacterial strains and water samples from China. Four Cylindrospermopsis strains and two Raphidiopsis strains were confirmed to produce CYNs. Mutant cyrI and cyrK genes were observed in these strains. Cloned cyr gene sequences from eight water bodies were clustered with cyr genes from Cylindrospermopsis and Raphidiopsis (C/R group) in the phylogenetic trees with high similarities (99%). Four cyrI sequence types and three cyrJ sequence types were observed to have different sequence insertions and repeats. Phylogenetic analysis of the rpoC1 sequences of the C/R group revealed four conserved clades, namely, clade I, clade II, clade III, and clade V. High sequence similarities (>97%) in each clade and a divergent clade IV were observed. Therefore, CYN producers were sporadically distributed in congeneric and paraphyletic C/R group species in Chinese freshwater ecosystems. In the evolution of cyr genes, intragenomic translocations and intergenomic transfer between local Cylindrospermopsis and Raphidiopsis were emphasized and probably mediated by transposases. This research confirms the existence of CYN-producing Cylindrospermopsis in China and reveals the distinctive variations of cyr genes.

Suppressing cyanobacterial dominance by UV-LED TiO2-photocatalysis in a drinking water reservoir: A mesocosm study.

Cyanobacteria and their toxic secondary metabolites present challenges for water treatment globally. In this study we have assessed TiO2 immobilized onto recycled foamed glass beads by a facile calcination method, combined in treatment units with 365 nm UV-LEDs. The treatment system was deployed in mesocosms within a eutrophic Brazilian drinking water reservoir. The treatment units were deployed for 7 days and suppressed cyanobacterial abundance by 85% while at the same time enhancing other water quality parameters; turbidity and transparency improved by 40 and 81% respectively. Genomic analysis of the microbiota in the treated mesocosms revealed that the composition of the cyanobacterial community was affected and the abundance of Bacteroidetes and Proteobacteria increased during cyanobacterial suppression. The effect of the treatment on zooplankton and other eukaryotes was also monitored. The abundance of zooplankton decreased while Chrysophyte and Alveolata loadings increased. The results of this proof-of-concept study demonstrate the potential for full-scale, in-reservoir application of advanced oxidation processes as complementary water treatment processes.

Degradation of Multiple Peptides by Microcystin-Degrader Paucibacter toxinivorans (2C20).

Since conventional drinking water treatments applied in different countries are inefficient at eliminating potentially toxic cyanobacterial peptides, a number of bacteria have been studied as an alternative to biological filters for the removal of microcystins (MCs). Here, we evaluated the degradation of not only MCs variants (-LR/DM-LR/-RR/-LF/-YR), but also non-MCs peptides (anabaenopeptins A/B, aerucyclamides A/D) by Paucibactertoxinivorans over 7 days. We also evaluated the degradation rate of MC-LR in a peptide mix, with all peptides tested, and in the presence of M. aeruginosa crude extract. Furthermore, biodegradation was assessed for non-cyanobacterial peptides with different chemical structures, such as cyclosporin A, (Glu1)-fibrinopeptide-B, leucine-enkephalin, and oxytocin. When cyanopeptides were individually added, P. toxinivorans degraded them (99%) over 7 days, except for MC-LR and -RR, which decreased by about 85 and 90%, respectively. The degradation rate of MC-LR decreased in the peptide mix compared to an individual compound, however, in the presence of the Microcystis extract, it was degraded considerably faster (3 days). It was noted that biodegradation rates decreased in the mix for all MCs while non-MCs peptides were immediately degraded. UPLC-QTOF-MS/MS allowed us to identify two linear biodegradation products for MC-LR and MC-YR, and one for MC-LF. Furthermore, P. toxinivorans demonstrated complete degradation of non-cyanobacterial peptides, with the exception of oxytocin, where around 50% remained after 7 days. Thus, although P. toxinivorans was previously identified as a MC-degrader, it also degrades a wide range of peptides under a range of conditions, which could be optimized as a potential biological tool for water treatment.

Ecophysiological Aspects and sxt Genes Expression Underlying Induced Chemical Defense in STX-Producing Raphidiopsis raciborskii (Cyanobacteria) against the Zooplankter Daphnia gessneri.

Cyanobacteria stand out among phytoplankton when they form massive blooms and produce toxins. Because cyanotoxin genes date to the origin of metazoans, the hypothesis that cyanotoxins function as a defense against herbivory is still debated. Although their primary cellular function might vary, these metabolites could have evolved as an anti-predator response. Here we evaluated the physiological and molecular responses of a saxitoxin-producing Raphidiopsis raciborskii to infochemicals released by the grazer Daphnia gessneri. Induced chemical defenses were evidenced in R. raciborskii as a significant increase in the transcription level of sxt genes, followed by an increase in saxitoxin content when exposed to predator cues. Moreover, cyanobacterial growth decreased, and no significant effects on photosynthesis or morphology were observed. Overall, the induced defense response was accompanied by a trade-off between toxin production and growth. These results shed light on the mechanisms underlying zooplankton-cyanobacteria interactions in aquatic food webs. The widespread occurrence of the cyanobacterium R. raciborskii in freshwater bodies has been attributed to its phenotypic plasticity. Assessing the potential of this species to thrive over interaction filters such as zooplankton grazing pressure can enhance our understanding of its adaptive success.

Effect of hydrogen peroxide on natural phytoplankton and bacterioplankton in a drinking water reservoir: Mesocosm-scale study.

Cyanobacterial blooms are increasingly reported worldwide, presenting a challenge to water treatment plants and concerning risks to human health and aquatic ecosystems. Advanced oxidative processes comprise efficient and safe methods for water treatment. Hydrogen peroxide (H2O2) has been proposed as a sustainable solution to mitigate bloom-forming cyanobacteria since this group presents a higher sensitivity compared to other phytoplankton, with no major risks to the environment at low concentrations. Here, we evaluated the effects of a single H2O2 addition (10 mg L-1) over 120 h in mesocosms introduced in a reservoir located in a semi-arid region presenting a Planktothrix-dominated cyanobacterial bloom. We followed changes in physical and chemical parameters and in the bacterioplankton composition. H2O2 efficiently suppressed cyanobacteria, green algae, and diatoms over 72 h, leading to an increase in transparency and dissolved organic carbon, and a decrease in dissolved oxygen and pH, while nutrient concentrations were not affected. After 120 h, cyanobacterial abundance remained low and green algae became dominant. 16S rRNA sequencing revealed that the original cyanobacterial bloom was composed by Planktothrix, Cyanobium and Microcystis. Only Cyanobium increased in relative abundance at 120 h, suggesting regrowth. A prominent change in the composition of heterotrophic bacteria was observed with Exiguobacterium, Paracoccus and Deinococcus becoming the most abundant genera after the H2O2 treatment. Our results indicate that this approach is efficient in suppressing cyanobacterial blooms and improving water quality in tropical environments. Monitoring changes in abiotic parameters and the relative abundance of specific bacterial taxa could be used to anticipate the regrowth of cyanobacteria after H2O2 degradation and to indicate where in the reservoir H2O2 should be applied so the effects are still felt in the water treatment plant intake.

Effects of saxitoxins exposure on oligodendrocyte development in mouse neonates.

Saxitoxins (STXs) are neurotoxins produced by cyanobacteria and dinoflagellates, and they are primarily known to block voltage-gated sodium channels in neurons. The present study aimed to obtain further information regarding the effects of these toxins on neurodevelopment by investigating the responses of murine subventricular zone (SVZ) neural progenitors to STXs. An in vitro neonatal mouse SVZ explant model was exposed to different concentrations of toxic cyanobacterial extracts to evaluate the migration and differentiation of SVZ-derived progenitor cells. To test the ability of STX to cross the placental barrier, pregnant mice received a single intraperitoneal injection of STXs (7.5 µg/kg body weight) on gestational day fifteen. Immunocytochemistry was performed to detect proliferating and differentiating progenitors, including oligodendrocyte progenitor cells (OPCs). It was found that specific proliferation of OPCs was significantly increased, but there was no corresponding increase in the number of differentiated oligodendrocytes, which may indicate a negative effect on the maturation process of these cells. Additionally, the data showed that STXs crossed the placental barrier. Thus, STXs can be considered a potential risk to fetal neurodevelopment.

Morphology and molecular phylogeny of a new PST-producing dinoflagellate species: Alexandrium fragae sp. nov. (Gonyaulacales, dinophyceae).

The genus Alexandrium comprises some of the most potentially toxic marine algae. A new toxic species of Alexandrium, A. fragae sp. nov., was found in Guanabara Bay, Rio de Janeiro, southern Brazil. The new species produces GTX2&3 and STX. The cell morphology of A. fragae resembles A. minutum in many characters, including the small size; the rounded-elliptical shape; and the shapes of the apical pore complex (APC), first apical plate (1'), sixth precingular plate (6″), and anterior and posterior sulcal plates (s.a. and s.p.). The main diagnostic characters of A. fragae are the ornamentation pattern, smooth epitheca and reticulated hypotheca, all of which were present in both natural populations and cultures. Phylogenies inferred from the ITS, LSU, and SSU rDNA of A. fragae showed that A. fragae clustered in a well-supported clade, distinct from other Alexandrium species. Morphology and molecular analyses based on ITS and LSU rDNA indicated that A. fragae strains and Alexandrium sp. from Japan (D163C5, D164C6) are a single species. Our findings suggest that the Alexandrium morphotype with a smooth epitheca and reticulated hypotheca, previously identified as A. minutum in different geographic regions, may corresponds to A. fragae.

Biomonitoring of cyanotoxins in two tropical reservoirs by cladoceran toxicity bioassays.

This study evaluates the potential for the use of cladocerans in biomonitoring of cyanobacterial toxins. Two zooplankton species (Daphnia gessneri and Moina micrura) were cultivated in the laboratory for use in acute (48 h) and chronic (10 days) bioassays. Water samples were collected from two reservoirs and diluted in mineral water at four concentrations. Survivorship in the acute bioassays was used to calculate LC50, and survivorship and fecundity in chronic bioassays were used to calculate the intrinsic population growth rate (r) and the EC50. Analysis of phytoplankton in the water samples from one reservoir revealed that cyanobacteria were the dominant group, represented by the genera Anabaena, Cylindrospermopsis, and Microcystis. Results of bioassays showed adverse effects including death, paralysis, and reduced population growth rate, generally proportional to the reservoir water concentration. These effects may be related to the presence of cyanobacteria toxins (microcystins or saxitoxins) in the water.

Intraspecific variability in response to phosphorus depleted conditions in the cyanobacteria Microcystis aeruginosa and Raphidiopsis raciborskii.

Phosphorus loading plays an important role in the occurrence of cyanobacterial blooms and understanding how this nutrient affects the physiology of cyanobacteria is imperative to manage these phenomena. Microcystis aeruginosa and Raphidiopsis raciborskii are cyanobacterial species that form potentially toxic blooms in freshwater ecosystems worldwide. Blooms comprise numerous strains with high trait variability, which can contribute to the widespread distribution of these species. Here, we explored the intraspecific variability in response to phosphorus depleted conditions (P-) testing five strains of each species. Strains could be differentiated by cell volume or genetic profiles except for those of the same species, sampling location and date, though these presented differences in their response to (P-). Although differently affected by (P-) over 10 days, all strains were able to grow and maintain photosynthetic activity. For most M. aeruginosa and R. raciborskii strains growth rates were not significantly different comparing (P+) and (P-) conditions. After ten days in (P-), only one M. aeruginosa strain and two R. raciborskii strains showed reduction in biovolume yield as compared to (P+) but in most strains chlorophyll-a concentrations were lower in (P-) than in (P+). Reduced photosystem II efficiency was found for only one R. raciborskii strain while all M. aeruginosa strains were affected. Only two M. aeruginosa and one R. raciborskii strain increased alkaline phosphatase activity under (P-) as compared to (P+). Variation in P-uptake was also observed but comparison among strains yielded homogeneous groups comprised of representatives of both species. Comparing the response of each species as a whole, the (P-) condition affected growth rate, biovolume yield and chlorophyll yield. However, these parameters revealed variation among strains of the same species to the extent that differences between M. aeruginosa and R. raciborskii were not significant. Taken together, these results do not support the idea that R. raciborskii, as a species, can withstand phosphorus limitation better than M. aeruginosa and also point that the level of intraspecific variation may preclude generalizations based on studies that use only one or few strains.

Planktonic microbial profiling in water samples from a Brazilian Amazonian reservoir.

Our comprehension of the dynamics and diversity of freshwater planktonic bacterial communities is far from complete concerning the Brazilian Amazonian region. Therefore, reference studies are urgently needed. We mapped bacterial communities present in the planktonic communities of a freshwater artificial reservoir located in the western Amazonian basin. Two samples were obtained from rainy and dry seasons, the periods during which water quality and plankton diversity undergo the most significant changes. Hypervariable 16S rRNA and shotgun sequencing were performed to describe the first reference of a microbial community in an Amazonian lentic system. Microbial composition consisted mainly of Betaproteobacteria, Cyanobacteria, Alphaproteobacteria, and Actinobacteria in the dry period. The bacteria distribution in the rainy period was notably absent of Cyanobacteria. Microcystis was observed in the dry period in which the gene cluster for cyanotoxins was found. Iron acquisition gene group was higher in the sample from the rainy season. This work mapped the first inventory of the planktonic microbial community of a large water reservoir in the Amazon, providing a reference for future functional studies and determining other communities and how they interact.

Close Link Between Harmful Cyanobacterial Dominance and Associated Bacterioplankton in a Tropical Eutrophic Reservoir.

Cyanobacteria tend to become the dominant phytoplankton component in eutrophic freshwater environments during warmer seasons. However, general observations of cyanobacterial adaptive advantages in these circumstances are insufficient to explain the prevalence of one species over another in a bloom period, which may be related to particular strategies and interactions with other components of the plankton community. In this study, we present an integrative view of a mixed cyanobacterial bloom occurring during a warm, rainy period in a tropical hydropower reservoir. We used high-throughput sequencing to follow temporal shifts in the dominance of cyanobacterial genera and shifts in the associated heterotrophic bacteria community. The bloom occurred during late spring-summer and included two distinct periods. The first period corresponded to Microcystis aeruginosa complex (MAC) dominance with a contribution from Dolichospermum circinale; this pattern coincided with high water retention time and low transparency. The second period corresponded to Cylindrospermopsis raciborskii and Synechococcus spp. dominance, and the reservoir presented lower water retention time and higher water transparency. The major bacterial phyla were primarily Cyanobacteria and Proteobacteria, followed by Actinobacteria, Bacteroidetes, Verrucomicrobia, and Planctomycetes. Temporal shifts in the dominance of cyanobacterial genera were not only associated with physical features of the water but also with shifts in the associated heterotrophic bacteria. The MAC bloom was associated with a high abundance of Bacteroidetes, particularly Cytophagales. In the second bloom period, Planctomycetes increased in relative abundance, five Planctomycetes OTUs were positively correlated with Synechococcus or C. raciborskii OTUs. Our results suggest specific interactions of the main cyanobacterial genera with certain groups of the heterotrophic bacterial community. Thus, considering biotic interactions may lead to a better understanding of the shifts in cyanobacterial dominance.

Oxidative imbalance in mice intoxicated by microcystin-LR can be minimized.

Microcystins-LR (MC-LR) is a cyanotoxin produced by cyanobacteria. We evaluated the antioxidant potential of LASSBio-596 (LB-596, inhibitor of phosphodiesterases 4 and 5), per os, and biochemical markers involved in lung and liver injury induced by exposure to sublethal dose of MC-LR. Fifty male Swiss mice received an intraperitoneal injection of 60 µL of saline (CTRL group, n = 20) or a sublethal dose of MC-LR (40 µg/kg, TOX group, n = 20). After 6 h the animals received either saline (TOX and CTRL groups) or LB-596 (50 mg/kg, TOX + LASS group, n = 10) by gavage. At 6 h after exposure, respiratory mechanics was evaluated in 10 CTRL and 10 TOX mice: there was a significant increase of all lung mechanics parameters (static elastance, viscoelastic component of elastance and lung resistive and viscoelastic/inhomogeneous pressures) in TOX compared to CTRL. 8 h after saline or MC-LR administration, i.e., 2 h after treatment with LB-596, blood serum levels of alanine aminotransferase and aspartate aminotransferase, activity of superoxide dismutase, catalase, and content of malondialdehyde and carbonyl in lung and liver, NADPH oxidase 2 and 4 mRNA expressions, dual oxidase enzyme activity and H2O2 generation were analyzed in lung homogenates. All parameters were significantly higher in TOX than in the other groups. There was no significant difference between CTRL and TOX + LASS. MC-LR deteriorated lung and liver functions and induced redox imbalance in them, which was prevented by oral administration of LB-596.

Effects of microcystin-LR on mouse lungs.

Toxic cyanobacteria blooms in drinking water supplies have been an increasing public health concern all over the world. Human populations can be exposed to microcystins, an important family of cyanotoxins, mainly by oral ingestion. However, inhalation from recreational water and hemodialysis can represent other routes. This study investigated changes in respiratory mechanics, histology, protein phosphatase (PP) 1 and 2A activity and microcystin in lung of adult mice injected intraperitoneally (i.p.) with microcystin-LR. Thirty-six mice were divided into control (CTRL) and test (CYANO) groups. CTRL group received an i.p. injection of saline and the CYANO group received 40 microg MCYST-LR/kg i.p. After 2 and 8 h, and 1, 2 and 4 days after toxin injection, six mice from each group were sampled for analyses. Resistive and viscoelastic pressures, static and dynamic elastances augmented at 2 h in CYANO and so remained until day 4. Alveolar collapse and inflammatory cell infiltration were found 2h after the injection, reaching peak values at 8 h. However, no microcystin or inhibition of PPases could be detected in mice lungs. In conclusion, MCYST-LR led to a rapid increase in lung impedance and an inflammatory response with interstitial edema and inflammatory cell recruitment in mice.

Mercury methylation and bacterial activity associated to tropical phytoplankton.

The methylated form of mercury (Hg), methylmercury (MeHg), is one of the most toxic pollutants. Biotic and/or abiotic methylation, often associated to sulfate-reducing bacteria metabolism, occurs in aquatic environments and in many tropical areas, mostly in the periphyton associated to floating macrophyte roots. Data about mercury methylation by phytoplankton are scarce and the aim of this study was to verify the biotic influence in the methylation process in Microcystis aeruginosa and Sineccocystis sp. laboratory strains and in natural populations of phytoplankton from two different aquatic systems, the mesotrophic Ribeirão das Lajes reservoir and hypereutrophic oligohaline Jacarepaguá lagoon, Rio de Janeiro state, Brazil. Adapted radiochemical techniques were used to measure sulfate-reduction, mercury methylation and bacterial activity in phytoplankton samples. Methyl-(203)Hg formation from added inorganic (203)Hg and (3)H-Leucine uptake were measured by liquid scintillation as well as sulfate-reduction, estimated as H(2)(35)S produced from added Na(2)(35)SO(4). There was no significant difference in low methylation potentials (0.37%) among the two cyanobacterium species studied in laboratory conditions. At Ribeirão das Lajes reservoir, there was no significant difference in methylation, bacterial activity and sulfate-reduction of surface sediment between the sampling points. Methylation in sediments (3-4%) was higher than in phytoplankton (1.5%), the opposite being true for bacterial activity (sediment mean 6.6 against 150.3 nmol gdw(-1) h(-1) for phytoplankton samples). At Jacarepaguá lagoon, an expressive bacterial activity (477.1 x 10(3) nmol gdw(-1) h(-1) at a concentration of 1000 nM leucine) and sulfate-reduction ( approximately 21% H(2)(35)S trapped) associated to phytoplankton (mostly cyanobacteria M. aeruginosa) was observed, but mercury methylation was not detected.

Is qPCR a Reliable Indicator of Cyanotoxin Risk in Freshwater?

The wide distribution of cyanobacteria in aquatic environments leads to the risk of water contamination by cyanotoxins, which generate environmental and public health issues. Measurements of cell densities or pigment contents allow both the early detection of cellular growth and bloom monitoring, but these methods are not sufficiently accurate to predict actual cyanobacterial risk. To quantify cyanotoxins, analytical methods are considered the gold standards, but they are laborious, expensive, time-consuming and available in a limited number of laboratories. In cyanobacterial species with toxic potential, cyanotoxin production is restricted to some strains, and blooms can contain varying proportions of both toxic and non-toxic cells, which are morphologically indistinguishable. The sequencing of cyanobacterial genomes led to the description of gene clusters responsible for cyanotoxin production, which paved the way for the use of these genes as targets for PCR and then quantitative PCR (qPCR). Thus, the quantification of cyanotoxin genes appeared as a new method for estimating the potential toxicity of blooms. This raises a question concerning whether qPCR-based methods would be a reliable indicator of toxin concentration in the environment. Here, we review studies that report the parallel detection of microcystin genes and microcystin concentrations in natural populations and also a smaller number of studies dedicated to cylindrospermopsin and saxitoxin. We discuss the possible issues associated with the contradictory findings reported to date, present methodological limitations and consider the use of qPCR as an indicator of cyanotoxin risk.

Understanding the winning strategies used by the bloom-forming cyanobacterium Cylindrospermopsis raciborskii.

The cyanobacterium Cylindrospermopsis raciborskii is a widespread species increasingly being recorded in freshwater systems around the world. It is of particular concern because strains in some geographic areas are capable of producing toxins with implications for human and animal health. Studies of this species have increased rapidly in the last two decades, especially in the southern hemisphere where toxic strains are prevalent. A clearer picture is emerging of the strategies adopted by this species to bloom and out-compete other species. This species has a high level of flexibility with respect to light and nutrients, with higher temperatures and carbon dioxide also promoting growth. There are two types of toxins produced by C. raciborskii: cylindrospermopsins (CYNs) and saxitoxins (STXs). The toxins CYNs are constitutively produced irrespective of environmental conditions and the ecological or physiological role is unclear, while STXs appear to serve as protection against high salinity and/or water hardness. It is also apparent that strains of this species can vary substantially in their physiological responses to environmental conditions, including CYNs production, and this may explain discrepancies in findings from studies in different geographical areas. The combination of a flexible strategy with respect to environmental conditions, and variability in strain response makes it a challenging species to manage. Our ability to improve bloom prediction will rely on a more detailed understanding of the complex physiology of this species.

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.

Repeated intranasal exposure to microcystin-LR affects lungs but not nasal epithelium in mice.

Microcystin-LR (MC-LR) is a harmful cyanotoxin able to induce adverse outcomes in the respiratory system. We aimed to examine the lungs and nasal epithelium of mice following a sub-chronic exposure to MC-LR. Swiss mice were intranasally instilled with 10 µL of distilled water (CTRL, n = 10) or 6.7 ng/kg of MC-LR diluted in 10 µL of distilled water (TOX, n = 8) during 30 consecutive days. Respiratory mechanics was measured in vivo and histology measurements (morphology and inflammation) were assessed in lungs and nasal epithelium samples 24 h after the last intranasal instillation. Despite the lack of changes in the nasal epithelium, TOX mice displayed an increased amount of PMN cells in the lungs (× 10(-3)/µm(2)), higher lung static elastance (cmH2O/mL), resistive and viscoelastic/inhomogeneous pressures (cmH2O) (7.87 ± 3.78, 33.96 ± 2.64, 1.03 ± 0.12, 1.01 ± 0.08, respectively) than CTRL (5.37 ± 4.02, 26.65 ± 1.24, 0.78 ± 0.06, 0.72 ± 0.05, respectively). Overall, our findings suggest that the nasal epithelium appears more resistant than lungs in this model of MC-LR intoxication.

Human intoxication by microcystins during renal dialysis treatment in Caruaru-Brazil.

In February 1996, an outbreak of illness occurred at a hemodialysis clinic in Caruaru, Pernambuco State-Brazil. At this clinic 116 (89%) of 131 patients experienced visual disturbances, nausea, vomiting, and muscle weakness, following routine haemodialysis treatment. Subsequently, 100 patients developed acute liver failure. As of December 1996, 52 of the deaths could be attributed to a common syndrome now called 'Caruaru Syndrome'. Examination of previous years' phytoplankton counts showed that cyanobacteria were dominant in the water supply reservoir since 1990. Analyses of carbon and other resins from the clinic's water treatment system plus serum and liver tissue of patients led to the identification of two groups of hepatotoxic cyanotoxins: microcystins (cyclic heptapeptides) in all of these samples and cylindrospermopsin (alkaloid hepatotoxic) in the carbon and resins. Comparison of victims symptoms and pathology with animal studies on these two cyanotoxins, leads us to conclude that the major contributing factor to death of the dialysis patients was intravenous exposure to microcystins, specifically microcystin-YR, -LR and -AR. In 2000, a review of the Brazilian regulation for drinking water quality, promoted by Brazilian Health Ministry with collaboration of PAHO, incorporated cyanobacteria and cyanotoxins into this new regulation as parameters that must to be monitored for water quality control.

Accumulation and depuration of microcystins (cyanobacteria hepatotoxins) in Tilapia rendalli (Cichlidae) under laboratory conditions.

In order to understand accumulation and depuration of microcystins (MCYSTs) in Tilapia rendalli, three experiments with juveniles were done. The experiments simulated the fish diet during a Microcystis aeruginosa bloom in three different situations. In the first one each fish received daily, during 15 days, fish food plus toxic cells of M. aeruginosa (20.4 microg MCYSTs fish(-1) day(-1)). In the following 15 days they were fed without toxic cells. In the second experiment, fish were fed only with toxic cells during 28 days (14.6 microg MCYSTs fish(-1) day(-1)) and in the third experiment, during 42 days, fish were fed with fish food plus toxic cells (29.2 microg MCYSTs fish(-1) day(-1)) previously disrupted (to simulate a senescent bloom). MCYSTs analyses were done by enzyme-linked immunosorbent assay (ELISA) in liver and muscle samples in all experiments and in faeces in the first one (only in the depuration period). The results demonstrated different profiles of MCYSTs accumulation in liver and muscle of T. rendalli. Comparing the experiments, the highest MCYSTs accumulation in the liver (2.8 microg g(-1)) occurred in the second one, where fish had only toxic cells as feeding source. In the first experiment, the highest MCYSTs accumulation in liver (0.6 microg MCYSTs g(-1)) was observed during the accumulation period, while in muscle, interestingly, the highest concentration (0.05 microg MCYSTs g(-1)) occurred in the depuration period. In this same period, it was also observed elimination of toxins through faeces. The second and third experiments showed almost the same average concentrations in tissues although fish have received more MCYSTs in third one. With respect to implications of the fish comsumption, MCYSTs accumulation in muscle of T. rendalli in all three experiments reached concentrations that would represent an intake of these toxins above the tolerable limit for humans and these results confirmed our previous observations from a field study. In conclusion, in this study it was observed that T. rendalli is able to accumulate MCYSTs and the availability of other feeding sources, besides toxic cells, probably interferes with the accumulation rate. Therefore, the occurrence of toxic cyanobacterial blooms produncing MCYSTs in aquaculture ponds could represent a risk to the quality of fish to the consumers.