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Ian R Falconer - One of the best experts on this subject based on the ideXlab platform.
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interaction of the Cyanobacterial Toxin cylindrospermopsin with the eukaryotic protein synthesis system
Toxicon, 2008Co-Authors: Suzanne Froscio, W. A. Wickramasinghe, G R Shaw, Andrew R Humpage, Ian R FalconerAbstract:The Cyanobacterial Toxin cylindrospermospin (CYN) was shown to inhibit the eukaryotic protein synthesis apparatus with similar potency in plant and mammalian cell extracts, IC50 of 334 nM in wheat germ extract and 110 nM in reticulocyte lysate. [14C] CYN binding was assessed in reticulocyte lysates, following separation of ribosomes from free Toxin on Sephadex G-50 size exclusion columns. While Toxin binding was shown to be concentration-dependent (60–600 nM [14C] CYN), it did not correlate with ribosome content. The molar ratio of Toxin to ribosomes was 0.02:1 at the binding plateau. Significant binding of [14C] CYN was identified in both the ribosome fraction (>90% total ribosomes) and the first wash fraction eluting from the Sephadex G-50 column, despite the low ribosome content ( 100 kDa. These results suggest that CYN's target may not be the ribosome itself, but rather one of the soluble proteins associated with the eukaryotic translation system.
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the Cyanobacterial Toxin cylindrospermopsin induces fetal toxicity in the mouse after exposure late in gestation
Toxicon, 2007Co-Authors: E H Rogers, Ian R Falconer, Andrew R Humpage, R D Zehr, M I Gage, M Marr, Neil ChernoffAbstract:Abstract Cylindrospermopsin (cyn) is a Cyanobacterial Toxin implicated in human and wildlife poisonings. We have completed studies investigating the potential of purified cyn to induce developmental toxicity in mammals. The teratology study involved intraperitoneal injections (8.0–128 μg kg −1 ) on gestational days (GD) 8–12 with subsequent examination of term fetuses for viability, weight and morphological anomalies. Cyn was lethal to a significant portion of the dams receiving ⩾32 μg kg −1 . Surviving pregnant females were killed and fetuses removed for examination. Analysis indicates no adverse effects on litter size, fetal weight, or incidence of anomalies. Subsequently, 50 μg kg −1 cyn was administered on GD 8–12 or 13–17. Animals were allowed to give birth and litters monitored for growth and viability. A reduction in litter size occurred in treated groups. Avg. pup wt. was only affected in the GD 13–17 group. GD 13–17 dams did not exhibit the toxicity noted in the GD 8–12 group but gave birth significantly earlier than controls. There was a significant number of dead GD 13–17 pups and incidences of blood in the gastrointestinal tract and hematomas in the tips of the tails in survivors. Pups were cross-fostered to control mothers in litters of 10. On postnatal days (PND) 5–6 there were no significant differences in weight gain or viability in GD 8–12 litters, while GD 13–17 litters had significantly reduced weight gain and viability. GD 13–17 exposed male pups still weighed significantly less than the controls after 15 months.
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cylindrospermopsin genotoxicity and cytotoxicity role of cytochrome p 450 and oxidative stress
Journal of Toxicology and Environmental Health, 2005Co-Authors: Andrew R Humpage, Suzanne Froscio, Philip Burcham, Frank Fontaine, Ian R FalconerAbstract:Cylindrospermopsin (CYN) is a Cyanobacterial Toxin found in drinking-water sources world wide. It was the likely cause of human poisonings in Australia and possibly Brazil. Although CYN itself is a potent protein synthesis inhibitor, its acute toxicity appears to be mediated by cytochrome p-450 (CYP450)-generated metabolites. CYN also induces genotoxic effects both in vitro and in vivo, and preliminary evidence suggests that tumors are generated by oral exposure to CYN. To understand the role of CYP450-activated CYN metabolites on in vitro genotoxicity, this study quantified the process in primary mouse hepatocytes using the COMET assay in both the presence and absence of CYP450 inhibitors known to block acute CYN cytotoxicity. CYN was cytotoxic at concentrations above 0.1 μM(EC50 = 0.5 μM) but produced significant increases in Comet tail length, area, and tail moment at 0.05 μM and above; hence genotoxicity is unlikely to be secondary to metabolic disruption due to toxicity. The CYP450 inhibitors omepraz...
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oral toxicity of the Cyanobacterial Toxin cylindrospermopsin in male swiss albino mice determination of no observed adverse effect level for deriving a drinking water guideline value
Environmental Toxicology, 2003Co-Authors: Andrew R Humpage, Ian R FalconerAbstract:The Cyanobacterial Toxin cylindrospermopsin (CYN) is a frequent contaminant of freshwaters throughout the world, including those that are sources of drinking water. The first cases of human poisoning attributed to this Toxin occurred from a treated drinking water supply in Queensland, Australia, in 1979. The Toxin causes extensive damage to the liver, kidneys, spleen, heart, and other organs. It is known to be a potent protein synthesis inhibitor, but there is mounting evidence for genotoxicity and that it metabolizes to even more toxic forms. As part of a risk assessment process leading to a guideline for a safe drinking water level for this Toxin, we performed a series of experiments to determine a no-observed-adverse-effect level (NOAEL) for this Toxin. In the first trial male mice were exposed to CYN-containing Cyanobacterial extract in their drinking water (0–657 μg CYN kg−1 day−1) for 10 weeks. In the second trial mice received purified CYN by daily gavage (0–240 μg CYN kg−1 day−1) for 11 weeks. Body and organ weights were recorded; urine, serum, and hematology analyses were performed; and histopathological examination of tissues was carried out. Body weights were significantly increased at low doses (30 and 60 μg kg−1 day−1) and decreased at high doses (432 and 657 μg kg−1 day−1). Liver and kidney weights were significantly increased at doses of 240 μg kg−1 day−1 and 60 μg kg−1 day−1, respectively. Serum bilirubin levels were significantly increased and bile acids significantly decreased at doses of 216 μg kg day−1 and greater. Urine total protein was significantly decreased at doses above 60 μg kg−1 day−1. The kidney appeared to be the more sensitive organ to this Toxin. If it is assumed that increased organ weights and changes in functional capacity are responses to an underlying toxic effect, then the NOAEL based on this data is 30 μg kg−1 day−1, which, with standard calculations and uncertainty factors, provides a proposed guideline safety value of 1 μg/L in drinking water. © 2003 Wiley Periodicals, Inc. Environ Toxicol 18: 94–103, 2003.
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oral toxicity of the Cyanobacterial Toxin cylindrospermopsin in male swiss albino mice determination of no observed adverse effect level for deriving a drinking water guideline value
Environmental Toxicology, 2003Co-Authors: Andrew R Humpage, Ian R FalconerAbstract:: The Cyanobacterial Toxin cylindrospermopsin (CYN) is a frequent contaminant of freshwaters throughout the world, including those that are sources of drinking water. The first cases of human poisoning attributed to this Toxin occurred from a treated drinking water supply in Queensland, Australia, in 1979. The Toxin causes extensive damage to the liver, kidneys, spleen, heart, and other organs. It is known to be a potent protein synthesis inhibitor, but there is mounting evidence for genotoxicity and that it metabolizes to even more toxic forms. As part of a risk assessment process leading to a guideline for a safe drinking water level for this Toxin, we performed a series of experiments to determine a no-observed-adverse-effect level (NOAEL) for this Toxin. In the first trial male mice were exposed to CYN-containing Cyanobacterial extract in their drinking water (0-657 microg CYN kg(-1) day(-1)) for 10 weeks. In the second trial mice received purified CYN by daily gavage (0-240 microg CYN kg(-1) day(-1)) for 11 weeks. Body and organ weights were recorded; urine, serum, and hematology analyses were performed; and histopathological examination of tissues was carried out. Body weights were significantly increased at low doses (30 and 60 microg kg(-1) day(-1)) and decreased at high doses (432 and 657 microg kg(-1) day(-1)). Liver and kidney weights were significantly increased at doses of 240 microg kg(-1) day(-1) and 60 microg kg(-1) day(-1), respectively. Serum bilirubin levels were significantly increased and bile acids significantly decreased at doses of 216 microg kg day(-1) and greater. Urine total protein was significantly decreased at doses above 60 microg kg(-1) day(-1). The kidney appeared to be the more sensitive organ to this Toxin. If it is assumed that increased organ weights and changes in functional capacity are responses to an underlying toxic effect, then the NOAEL based on this data is 30 microg kg(-1) day(-1), which, with standard calculations and uncertainty factors, provides a proposed guideline safety value of 1 microg/L in drinking water.
Andrew R Humpage - One of the best experts on this subject based on the ideXlab platform.
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Cytotoxicity Screening for the Cyanobacterial Toxin Cylindrospermopsin
Journal of Toxicology and Environmental Health, 2009Co-Authors: Suzanne Froscio, Stella Fanok, Andrew R HumpageAbstract:The cell lines C3A, HepG2, NCI-87, HCT-8, HuTu-80, Caco-2, and Vero were screened for sensitivity to the Cyanobacterial Toxin cylindrospermopsin (CYN), with the aim of determining the most sensitive cells to be used in cytotoxicity tests. Cell lines were chosen to be representative of the organs targeted by the Toxin; liver, kidney, intestine, and were expected to have different metabolic activities and uptake capabilities. Over the range of cell lines tested, IC50 determinations at 24 h (MTT assay) ranged fourfold, from 1.5 μM for hepatocyte-derived cell lines (C3A IC50 = 1.5 ± 0.54; HepG2 IC50 = 1.5 ± 0.87) to 6.5 ± 3.3μM for the colon-derived Caco-2 cell line. The cell-line sensitivity seemed to decrease in cell lines derived from progressively more distal regions of the gastrointestinal tract: gastric > duodenal > ileal > colonic. The greater sensitivity of the hepatic cell lines to CYN was also apparent in 7-d exposure studies, with low Toxin concentrations exerting cytotoxic effects that were not se...
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interaction of the Cyanobacterial Toxin cylindrospermopsin with the eukaryotic protein synthesis system
Toxicon, 2008Co-Authors: Suzanne Froscio, W. A. Wickramasinghe, G R Shaw, Andrew R Humpage, Ian R FalconerAbstract:The Cyanobacterial Toxin cylindrospermospin (CYN) was shown to inhibit the eukaryotic protein synthesis apparatus with similar potency in plant and mammalian cell extracts, IC50 of 334 nM in wheat germ extract and 110 nM in reticulocyte lysate. [14C] CYN binding was assessed in reticulocyte lysates, following separation of ribosomes from free Toxin on Sephadex G-50 size exclusion columns. While Toxin binding was shown to be concentration-dependent (60–600 nM [14C] CYN), it did not correlate with ribosome content. The molar ratio of Toxin to ribosomes was 0.02:1 at the binding plateau. Significant binding of [14C] CYN was identified in both the ribosome fraction (>90% total ribosomes) and the first wash fraction eluting from the Sephadex G-50 column, despite the low ribosome content ( 100 kDa. These results suggest that CYN's target may not be the ribosome itself, but rather one of the soluble proteins associated with the eukaryotic translation system.
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the Cyanobacterial Toxin cylindrospermopsin induces fetal toxicity in the mouse after exposure late in gestation
Toxicon, 2007Co-Authors: E H Rogers, Ian R Falconer, Andrew R Humpage, R D Zehr, M I Gage, M Marr, Neil ChernoffAbstract:Abstract Cylindrospermopsin (cyn) is a Cyanobacterial Toxin implicated in human and wildlife poisonings. We have completed studies investigating the potential of purified cyn to induce developmental toxicity in mammals. The teratology study involved intraperitoneal injections (8.0–128 μg kg −1 ) on gestational days (GD) 8–12 with subsequent examination of term fetuses for viability, weight and morphological anomalies. Cyn was lethal to a significant portion of the dams receiving ⩾32 μg kg −1 . Surviving pregnant females were killed and fetuses removed for examination. Analysis indicates no adverse effects on litter size, fetal weight, or incidence of anomalies. Subsequently, 50 μg kg −1 cyn was administered on GD 8–12 or 13–17. Animals were allowed to give birth and litters monitored for growth and viability. A reduction in litter size occurred in treated groups. Avg. pup wt. was only affected in the GD 13–17 group. GD 13–17 dams did not exhibit the toxicity noted in the GD 8–12 group but gave birth significantly earlier than controls. There was a significant number of dead GD 13–17 pups and incidences of blood in the gastrointestinal tract and hematomas in the tips of the tails in survivors. Pups were cross-fostered to control mothers in litters of 10. On postnatal days (PND) 5–6 there were no significant differences in weight gain or viability in GD 8–12 litters, while GD 13–17 litters had significantly reduced weight gain and viability. GD 13–17 exposed male pups still weighed significantly less than the controls after 15 months.
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cylindrospermopsin genotoxicity and cytotoxicity role of cytochrome p 450 and oxidative stress
Journal of Toxicology and Environmental Health, 2005Co-Authors: Andrew R Humpage, Suzanne Froscio, Philip Burcham, Frank Fontaine, Ian R FalconerAbstract:Cylindrospermopsin (CYN) is a Cyanobacterial Toxin found in drinking-water sources world wide. It was the likely cause of human poisonings in Australia and possibly Brazil. Although CYN itself is a potent protein synthesis inhibitor, its acute toxicity appears to be mediated by cytochrome p-450 (CYP450)-generated metabolites. CYN also induces genotoxic effects both in vitro and in vivo, and preliminary evidence suggests that tumors are generated by oral exposure to CYN. To understand the role of CYP450-activated CYN metabolites on in vitro genotoxicity, this study quantified the process in primary mouse hepatocytes using the COMET assay in both the presence and absence of CYP450 inhibitors known to block acute CYN cytotoxicity. CYN was cytotoxic at concentrations above 0.1 μM(EC50 = 0.5 μM) but produced significant increases in Comet tail length, area, and tail moment at 0.05 μM and above; hence genotoxicity is unlikely to be secondary to metabolic disruption due to toxicity. The CYP450 inhibitors omepraz...
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oral toxicity of the Cyanobacterial Toxin cylindrospermopsin in male swiss albino mice determination of no observed adverse effect level for deriving a drinking water guideline value
Environmental Toxicology, 2003Co-Authors: Andrew R Humpage, Ian R FalconerAbstract:The Cyanobacterial Toxin cylindrospermopsin (CYN) is a frequent contaminant of freshwaters throughout the world, including those that are sources of drinking water. The first cases of human poisoning attributed to this Toxin occurred from a treated drinking water supply in Queensland, Australia, in 1979. The Toxin causes extensive damage to the liver, kidneys, spleen, heart, and other organs. It is known to be a potent protein synthesis inhibitor, but there is mounting evidence for genotoxicity and that it metabolizes to even more toxic forms. As part of a risk assessment process leading to a guideline for a safe drinking water level for this Toxin, we performed a series of experiments to determine a no-observed-adverse-effect level (NOAEL) for this Toxin. In the first trial male mice were exposed to CYN-containing Cyanobacterial extract in their drinking water (0–657 μg CYN kg−1 day−1) for 10 weeks. In the second trial mice received purified CYN by daily gavage (0–240 μg CYN kg−1 day−1) for 11 weeks. Body and organ weights were recorded; urine, serum, and hematology analyses were performed; and histopathological examination of tissues was carried out. Body weights were significantly increased at low doses (30 and 60 μg kg−1 day−1) and decreased at high doses (432 and 657 μg kg−1 day−1). Liver and kidney weights were significantly increased at doses of 240 μg kg−1 day−1 and 60 μg kg−1 day−1, respectively. Serum bilirubin levels were significantly increased and bile acids significantly decreased at doses of 216 μg kg day−1 and greater. Urine total protein was significantly decreased at doses above 60 μg kg−1 day−1. The kidney appeared to be the more sensitive organ to this Toxin. If it is assumed that increased organ weights and changes in functional capacity are responses to an underlying toxic effect, then the NOAEL based on this data is 30 μg kg−1 day−1, which, with standard calculations and uncertainty factors, provides a proposed guideline safety value of 1 μg/L in drinking water. © 2003 Wiley Periodicals, Inc. Environ Toxicol 18: 94–103, 2003.
Suzanne Froscio - One of the best experts on this subject based on the ideXlab platform.
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limited uptake of the Cyanobacterial Toxin cylindrospermopsin by vero cells
Toxicon, 2009Co-Authors: Suzanne Froscio, E Cannon, A R HumpageAbstract:Abstract Cylindrospermopsin (CYN) is a Cyanobacterial Toxin increasingly found in drinking water sources worldwide. Toxicity studies have shown CYN can induce effects in a range of different cell types with primary hepatocytes consistently shown to be the most sensitive cellular model. How CYN enters the intracellular environment is not clear, although the size and hydrophilic nature of the Toxin suggest it would not readily cross a lipid bilayer. In this study, a Vero cell line expressing green fluorescent protein (GFP) was used to monitor for CYN uptake based on the Toxin's potent effects on protein synthesis. Effects on the GFP signal were compared with inhibitors cycloheximide (CHEX) and emetine. While CYN potency was demonstrated in a cell-free system (CYN > CHEX > emetine) it was considerably reduced in the Vero-GFP cell model (CHEX, emetine > > CYN). In contrast to other inhibitors, CYN effects on GFP signal increased 6 fold over 4–24 h incubation indicating slow, progressive uptake of the Toxin. Confirming that the uptake process is not energy dependent CYN entry also occurred at 4 °C, while competition experiments excluded the uracil nucleobase transporter system as potential mechanism for CYN uptake. Dilution of media enhanced CYN uptake by Vero-GFP cells although mechanism by which this occurred is unknown.
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Cytotoxicity Screening for the Cyanobacterial Toxin Cylindrospermopsin
Journal of Toxicology and Environmental Health, 2009Co-Authors: Suzanne Froscio, Stella Fanok, Andrew R HumpageAbstract:The cell lines C3A, HepG2, NCI-87, HCT-8, HuTu-80, Caco-2, and Vero were screened for sensitivity to the Cyanobacterial Toxin cylindrospermopsin (CYN), with the aim of determining the most sensitive cells to be used in cytotoxicity tests. Cell lines were chosen to be representative of the organs targeted by the Toxin; liver, kidney, intestine, and were expected to have different metabolic activities and uptake capabilities. Over the range of cell lines tested, IC50 determinations at 24 h (MTT assay) ranged fourfold, from 1.5 μM for hepatocyte-derived cell lines (C3A IC50 = 1.5 ± 0.54; HepG2 IC50 = 1.5 ± 0.87) to 6.5 ± 3.3μM for the colon-derived Caco-2 cell line. The cell-line sensitivity seemed to decrease in cell lines derived from progressively more distal regions of the gastrointestinal tract: gastric > duodenal > ileal > colonic. The greater sensitivity of the hepatic cell lines to CYN was also apparent in 7-d exposure studies, with low Toxin concentrations exerting cytotoxic effects that were not se...
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interaction of the Cyanobacterial Toxin cylindrospermopsin with the eukaryotic protein synthesis system
Toxicon, 2008Co-Authors: Suzanne Froscio, W. A. Wickramasinghe, G R Shaw, Andrew R Humpage, Ian R FalconerAbstract:The Cyanobacterial Toxin cylindrospermospin (CYN) was shown to inhibit the eukaryotic protein synthesis apparatus with similar potency in plant and mammalian cell extracts, IC50 of 334 nM in wheat germ extract and 110 nM in reticulocyte lysate. [14C] CYN binding was assessed in reticulocyte lysates, following separation of ribosomes from free Toxin on Sephadex G-50 size exclusion columns. While Toxin binding was shown to be concentration-dependent (60–600 nM [14C] CYN), it did not correlate with ribosome content. The molar ratio of Toxin to ribosomes was 0.02:1 at the binding plateau. Significant binding of [14C] CYN was identified in both the ribosome fraction (>90% total ribosomes) and the first wash fraction eluting from the Sephadex G-50 column, despite the low ribosome content ( 100 kDa. These results suggest that CYN's target may not be the ribosome itself, but rather one of the soluble proteins associated with the eukaryotic translation system.
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cylindrospermopsin genotoxicity and cytotoxicity role of cytochrome p 450 and oxidative stress
Journal of Toxicology and Environmental Health, 2005Co-Authors: Andrew R Humpage, Suzanne Froscio, Philip Burcham, Frank Fontaine, Ian R FalconerAbstract:Cylindrospermopsin (CYN) is a Cyanobacterial Toxin found in drinking-water sources world wide. It was the likely cause of human poisonings in Australia and possibly Brazil. Although CYN itself is a potent protein synthesis inhibitor, its acute toxicity appears to be mediated by cytochrome p-450 (CYP450)-generated metabolites. CYN also induces genotoxic effects both in vitro and in vivo, and preliminary evidence suggests that tumors are generated by oral exposure to CYN. To understand the role of CYP450-activated CYN metabolites on in vitro genotoxicity, this study quantified the process in primary mouse hepatocytes using the COMET assay in both the presence and absence of CYP450 inhibitors known to block acute CYN cytotoxicity. CYN was cytotoxic at concentrations above 0.1 μM(EC50 = 0.5 μM) but produced significant increases in Comet tail length, area, and tail moment at 0.05 μM and above; hence genotoxicity is unlikely to be secondary to metabolic disruption due to toxicity. The CYP450 inhibitors omepraz...
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cell free protein synthesis inhibition assay for the Cyanobacterial Toxin cylindrospermopsin
Environmental Toxicology, 2001Co-Authors: Suzanne Froscio, Andrew R Humpage, Philip Burcham, Ian R FalconerAbstract:The Cyanobacterial Toxin cylindrospermopsin (CYN) is known to be a potent inhibitor of protein synthesis. This paper describes the use of a rabbit reticulocyte lysate translation system as a protein synthesis inhibition assay for CYN. A dose response curve for protein synthesis inhibition by CYN was constructed and was modeled to a sigmoidal dose response curve with variable slope (R2=0.98). In this assay, CYN has an IC50 of 120 nM [95% chemical interaction (CI)=111−130 nM] with a detection limit in the region of 50 nM in the assay solution. Application of the assay allows quantification of Toxin samples within the range 0.5–3.0 μM (200–1200 μg/L) CYN. To assess the usefulness of this assay, a range of toxic and nontoxic Cylindrospermopsis raciborskii extracts, including both laboratory strains and environmental samples, were assayed by protein synthesis inhibition. These CYN quantifications were then compared to quantifications obtained by high performance liquid chromatography (HPLC) and HPLC-tandem mass spectrometry (HPLCMS–MS). The results demonstrate that the protein synthesis inhibition assay correlates well with both HPLCMS–MS (r2=0.99) and HPLC (r2=0.97) quantifications. We conclude that this is an accurate and rapid assay for the measurement of cylindrospermopsin in Cyanobacterial extracts. © 2001 John Wiley & Sons, Inc. Environ Toxicol 16: 408–412, 2001
Stephan Pflugmacher - One of the best experts on this subject based on the ideXlab platform.
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translocation of the Cyanobacterial Toxin microcystin lr into guttation drops of triticum aestivum and remaining toxicity
Environmental Pollution, 2019Co-Authors: Stephan Pflugmacher, Amalia Sulk, Maranda EsterhuizenlondtAbstract:Abstract Uptake of the commonly occurring Cyanobacterial Toxin microcystin-LR (MC-LR) into crop plants via spray irrigation has been demonstrated. As other hazardous compounds such as pesticides were shown to be transported within plants, it was essential to understand the transport and fate of MC-LR in plants and the risks posed to grazers and other consumers. Of specific interest was to investigate if MC-LR could be detected in guttation drops and the toxicity thereof. Triticum aestivum (wheat) seedlings were exposed to 100 μg L−1 MC-LR in two separate experiments during which guttation drops were collected at various time points. The plants of one experiment were sectioned to investigate MC-LR distribution to the various plant appendages via liquid chromatography-tandem mass spectrometry analysis. After exposure, MC-LR could be detected in the roots, stems, leaves, and the guttation drops. However, the guttation drops were not toxic to Daphnia. As the environmentally relevant Toxin concentration used was not sufficient to promote mortality in Daphnia, the physiological effect in insects, which rely on guttation drops as a water source, remains unknown. Combined with other contaminants that insects may be exposed to, the additional MC-LR exposure could contribute to the overall toxicity through the “tears of death”.
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still challenging the ecological function of the Cyanobacterial Toxin microcystin what we know so far
Toxin Reviews, 2018Co-Authors: Azam Omidi, Maranda Esterhuizenlondt, Stephan PflugmacherAbstract:AbstractMicrocystins (MCs) are the most commonly studied cyanoToxins. While these past studies have mainly focused on the toxicity of MCs, the evolutionary history of life has shown that toxicity can be considered as an assigned role to MCs. Nowadays, there is a growing interest in understanding the importance of cyanoToxins in any of the physiological processes or beyond at the ecological level. This review evaluates variously proposed intracellular and extracellular functions of MCs and how they benefit the producing cyanobacterium. However, the strain-specific and divergent laboratory and field results obtained to date have made it difficult to generalize.
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potential role of engineered nanoparticles as contaminant carriers in aquatic ecosystems estimating sorption processes of the Cyanobacterial Toxin microcystin lr by tio2 nanoparticles
Colloids and Surfaces A: Physicochemical and Engineering Aspects, 2015Co-Authors: Annette Okupnik, Valeska Contardojara, Stephan PflugmacherAbstract:Abstract In the present study, the adsorption of a Cyanobacterial Toxin, microcystin-LR (MC-LR), onto titanium dioxide nanoparticles (TiO 2 -NPs) was investigated in aqueous solution. TiO 2 -NPs with different crystalline phases (anatase, rutile, anatase-rutile mixture) and MC-LR in an environmental relevant concentration for German water bodies were used to make results more applicable to environmental conditions. To investigate the adsorption mechanism as well as to determine the rate-limiting step of adsorption, analysis of the adsorption kinetic was performed and common kinetic models were applied to the experimental data. Additionally, the influence of nano-TiO 2 dosage (0.01–10 mg/L) and particle size on adsorption capacity was examined, the latter by comparing the adsorption data of TiO 2 -NPs to a bulk sized counterpart. The analysis of the adsorption kinetic indicates a complex adsorption mechanism and the involvement of pseudo-second-order chemisorption in the adsorption process. While only anatase TiO 2 showed a variation of MC-LR adsorption at different adsorbent concentrations, the particle size of the adsorbents appears to be the most influencing factor on Toxin adsorption. All TiO 2 -NPs exhibited higher adsorption capacities compared to the bulk TiO 2 counterpart, which was statistically significant for the environmental relevant concentration levels of TiO 2 -NPs. The obtained results emphasise the potential of TiO 2 -NPs to act as carriers for environmental contaminants such as the Cyanobacterial Toxin MC-LR, and thus to influence its fate in aquatic ecoystems.
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the fate of the Cyanobacterial Toxin β n methylamino l alanine in freshwater mussels
Ecotoxicology and Environmental Safety, 2014Co-Authors: Simone Downing, Stephan Pflugmacher, Valeska Contardojara, T G DowningAbstract:Abstract The Cyanobacterial neuroToxin, β- N -methylamino- l -alanine (BMAA) has been suggested as a causative agent for certain neurodegenerative diseases. This cyanoToxin bioaccumulates in an array of aquatic organisms, in which it occurs as both a free amino acid and in a protein-associated form. This study was intended to investigate the environmental fate of BMAA by examining the metabolism of isotopically labeled BMAA in four freshwater mussel species. All species showed substantial uptake of BMAA from the culture media. Data showed no significant evidence for BMAA catabolism in any of the animals but did suggest metabolism via the reversible covalent modification of BMAA in freshwater mussels, a process that appears to be variable in different species.
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promotion of oxidative stress in the aquatic macrophyte ceratophyllum demersum during biotransformation of the Cyanobacterial Toxin microcystin lr
Aquatic Toxicology, 2004Co-Authors: Stephan PflugmacherAbstract:Abstract Cyanobacterial Toxins have been shown to have adverse effects on mammals, birds and fish and are therefore being increasingly recognised as a potent stress and health hazard factor in aquatic ecosystems. Microcystins, which are cyclic heptapeptides and a main group of the cyanoToxins, are mainly retained within the producer-cells during Cyanobacterial bloom development. However, these Toxins are released into the surrounding medium by senescence and lysis of the blooms. The released Toxins could then come into contact with a wide range of aquatic organisms including invertebrates, fish and aquatic plants. In many organisms, biotransformation of the Toxins will take place via several glutathione-related conjugate. During the biotransformation process in which the Toxin and the Toxin conjugate are broken down, the formation of reactive oxygen species might occur. These reactive oxygen species activate several antioxidant enzymes such as superoxide dismutase, catalase, ascorbate peroxidase and also influence the glutathione-ascorbate cycle. Aim of this study was to investigate formation of the glutathione-conjugate, activation of glutathione S-transferases and the elevation of several antioxidant enzymes giving evidence for the promotion of oxidative stress by microcystins. During exposure of Ceratophyllum demersum to the Cyanobacterial Toxin microcystin-LR in an concentration of 5.0 μg/L, an elevation of microsomal and cytosolic glutathione S-transferase was measured, showing the beginning formation of the glutathione-Toxin conjugate. The superoxide dismutase as well as in parallel the hydrogen peroxide level increased giving evidence for oxidative stress in the aquatic plant. Other reactive oxygen detoxifiying enzymes were also elevated and the glutathione pool, expressed in reduced glutathione and glutathione disulfide concentration was changed accordingly.
Brett A Neilan - One of the best experts on this subject based on the ideXlab platform.
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directing the heterologous production of specific Cyanobacterial Toxin variants
ACS Chemical Biology, 2017Co-Authors: Rabia Mazmouz, Sarah E Ongley, Rocky Chau, Russell Pickford, Jason N Woodhouse, Brett A NeilanAbstract:Microcystins are globally the most commonly occurring freshwater cyanoToxins, causing acute poisoning and chronically inducing hepatocellular carcinoma. However, the detection and toxicological study of microcystins is hampered by the limited availability and high cost of pure Toxin standards. Biosynthesis of microcystin variants in a fast-growing heterologous host offers a promising method of achieving reliable and economically viable alternative to isolating Toxin from slow-growing Cyanobacterial cultures. Here, we report the heterologous expression of recombinant microcystin synthetases in Escherichia coli to produce [d-Asp3]microcystin-LR and microcystin-LR. We assembled a 55 kb hybrid polyketide synthase/nonribosomal peptide synthetase gene cluster from Microcystis aeruginosa PCC 7806 using Red/ET recombineering and replaced the native promoters with an inducible PtetO promoter to yield microcystin titers superior to M. aeruginosa. The expression platform described herein can be tailored to heterolog...
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Nodularin, a Cyanobacterial Toxin, is synthesized in planta by symbiotic Nostoc sp.
The ISME Journal, 2012Co-Authors: Michelle M Gehringer, Troco Kaan Mihali, Michelle C Moffitt, Lewis Adler, Alexandra A Roberts, Toby J T Mills, Claus Fieker, Brett A NeilanAbstract:The nitrogen-fixing bacterium, Nostoc , is a commonly occurring cyanobacterium often found in symbiotic associations. We investigated the potential of cycad Cyanobacterial endosymbionts to synthesize microcystin/nodularin. Endosymbiont DNA was screened for the aminotransferase domain of the Toxin biosynthesis gene clusters. Five endosymbionts carrying the gene were screened for bioactivity. Extracts of two isolates inhibited protein phosphatase 2A and were further analyzed using electrospray ionization mass spectrometry (ESI-MS)/MS. Nostoc sp. ‘ Macrozamia riedlei 65.1’ and Nostoc sp. ‘ Macrozamia serpentina 73.1’ both contained nodularin. High performance liquid chromatography (HPLC) HESI-MS/MS analysis confirmed the presence of nodularin at 9.55±2.4 ng μg−1 chlorophyll a in Nostoc sp. ‘ Macrozamia riedlei 65.1’ and 12.5±8.4 ng μg−1 Chl a in Nostoc sp. ‘ Macrozamia serpentina 73.1’ extracts. Further scans indicated the presence of the rare isoform [L-Har^2] nodularin, which contains L -homoarginine instead of L -arginine. Nodularin was also present at 1.34±0.74 ng ml^−1 (approximately 3 pmol per g plant ww) in the methanol root extracts of M. riedlei MZ65, while the presence of [L-Har^2] nodularin in the roots of M. serpentina MZ73 was suggested by HPLC HESI-MS/MS analysis. The ndaA - B and ndaF genomic regions were sequenced to confirm the presence of the hybrid polyketide/non-ribosomal gene cluster. A seven amino-acid insertion into the NdaA-C1 domain of N. spumigena NSOR10 protein was observed in all endosymbiont-derived sequences, suggesting the transfer of the nda cluster from N. spumigena to terrestrial Nostoc species. This study demonstrates the synthesis of nodularin and [L-Har^2] nodularin in a non- Nodularia species and the production of Cyanobacterial hepatoToxin by a symbiont in planta.
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characterization of the gene cluster responsible for cylindrospermopsin biosynthesis
Applied and Environmental Microbiology, 2008Co-Authors: Troco Kaan Mihali, Ralf Kellmann, Julia Muenchhoff, Kevin D Barrow, Brett A NeilanAbstract:Toxic Cyanobacterial blooms cause economic losses and pose significant public health threats on a global scale. Characterization of the gene cluster for the biosynthesis of the Cyanobacterial Toxin cylindrospermopsin (cyr) in Cylindrospermopsis raciborskii AWT205 is described, and the complete biosynthetic pathway is proposed. The cyr gene cluster spans 43 kb and is comprised of 15 open reading frames containing genes required for the biosynthesis, regulation, and export of the Toxin. Biosynthesis is initiated via an amidinotransfer onto glycine followed by five polyketide extensions and subsequent reductions, and rings are formed via Michael additions in a stepwise manner. The uracil ring is formed by a novel pyrimidine biosynthesis mechanism and tailoring reactions, including sulfation and hydroxylation that complete biosynthesis. These findings enable the design of toxic strain-specific probes and allow the future study of the regulation and biological role of cylindrospermopsin.
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molecular detection of genes responsible for Cyanobacterial Toxin production in the genera microcystis nodularia and cylindrospermopsis
Methods of Molecular Biology, 2004Co-Authors: Brendan P Burns, Martin L Saker, Michelle C Moffitt, Brett A NeilanAbstract:: Cyanobacteria are ubiquitous in the freshwater environment. Their success as a group in a wide range of aquatic habitats has been attributed to their unique physiological characteristics and their high adaptive ability over a wide range of environmental conditions. They are capable of reaching very high biomass levels, often dominating the other aquatic biota, and under some circumstances can accumulate near the water surface, producing scums. Such Cyanobacterial "blooms" are of particular concern in reservoirs used to supply potable water. Dense aggregations of Cyanobacterial cells may block water filters, and many species produce compounds that affect the taste and odor of water supplies. Of greatest concern, however, is the potential of many bloom-forming cyanobacteria to produce a wide range of toxic substances. These natural compounds, known as cyanoToxins, are chemically diverse and are usually either neuro- or hepatotoxic in pathology.
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absence of detectable levels of the Cyanobacterial Toxin microcystin lr carry over into milk
Toxicon, 2002Co-Authors: Andrew J Feitz, Tredwell Lukondeh, Michelle C Moffitt, Brendan P Burns, Daya Naidoo, Jim Della Vedova, James M Gooden, Brett A NeilanAbstract:: The potential for the carry-over of the Cyanobacterial Toxin, microcystin-LR, from feed to milk was assessed using four Holstein-Friesian cows in a 4 week feeding trial. Two cows were used as control and the other two dosed daily at increasing weekly concentrations of microcystins from zero to a maximum dosage of 13 microg Toxin kg x (-1) d x (-1) (or 7.4 mg Toxin day(-1)). The absence of any deviation from the control in terms of physiological response and plasma indicators (total bilirubin, gamma-glutamyl transpeptidase and alkaline phosphatase) suggests that the microcystin-LR dosage did not have a detrimental effect on cattle liver function or milk yield during the course of the study. While the milk production did decrease over the period of the trial, no difference was observed between control and dosed cattle. Protein phosphatase inhibition assays were successfully used to determine the presence of microcystin-LR in prepared milk samples with an average recovery of 88% for samples spiked with 0.6 microg x l(-1) microcystin-LR. The level of microcystin-LR in all milk samples during the trial was less than 0.2 microg x l(-1). This suggests that after digestion, microcystin--LR is either not present in milk or sufficiently modified to render it non-toxic.
