Toxicological comparison of diverse Cylindrospermopsis raciborskii strains: evidence of liver damage caused by a French C raciborskii strain

The freshwater cyanobacterium Cylindrospermopsis raciborskii is known to produce toxic effects in several countries. Acute and chronic exposures to C. raciborskii in Australia have been linked to liver damage (hepatotoxicity) with concomitant effects on the kidneys, adrenal glands, small intestine, lungs, thymus, and heart. The alkaloid cylindrospermopsin, which produces these toxic effects, is thought to be a potent inhibitor of protein synthesis. C. raciborskii strains producing cylindrospermopsin or analogue alkaloids have also been reported in Florida, USA, and Thailand. Brazilian isolates of C. raciborskii are also toxic but act by a different mechanism, causing acute death in mice with neurotoxic symptoms similar to those induced by the saxitoxins. In this article we compare the toxicity in the mouse of a C. raciborskii French strain with C. raciborskii strains from various other sources (Australia, Brazil, Mexico, and Hungary). We tested the toxicity of cell extracts by a mouse bioassay. Acute, fatal neurotoxicity was produced by the Brazilian strain, which was confirmed by liquid chromatography with fluorescence detection of the cell extracts, which revealed the presence of saxitoxin, neosaxitoxin, and decarbamoylsaxitoxin, along with two unidentified compounds. Acute hepatotoxicity with severe liver, kidney, and thymus damage was observed with the Australian cylindrospermopsin-producing strain. The Mexican and Hungarian strains were not found to be toxic to mice in our experimental conditions. No animals died after exposure to the extracts of the French C. raciborskii strain. Histological examination of the liver revealed moderate, multifocal necrosis characterized by small areas of hepatocellular necrosis, combined with disorganization of the parenchyma and congestion of the inner sinusoid. These symptoms and lesions resembled those induced by cylindrospermopsin, but the chemical analysis performed by liquid chromatography coupled with either a diode array detector or a mass spectrometer demonstrated that this toxin was not present in our culture extract.     

A comparative study of Florida strains of Cylindrospermopsis and Aphanizomenon for cylindrospermopsin production.

The toxin cylindrospermopsin (CYN) is produced by a variety of cyanobacterial genera. One of these, Cylindrospermopsis raciborskii, is generally assumed to be the source of CYN in lakes and rivers in Florida, USA. However, in this study, none of the eight Florida isolates of this species tested contained the genetic determinants involved in toxin production nor did they produce CYN. We show for the first time that Aphanizomenon ovalisporum isolated from a pond in this state has the genes putatively associated with CYN production. Analysis by liquid chromatography with mass spectrometric detection (LC/MS) revealed that it produced CYN in the range of 7.39-9.33 microg mg(-1) freeze-dried cells. 16S rDNA sequences of this strain showed 99.6% and 99.9% identity to published A. ovalisporum and Anabaena bergii 16S sequences, respectively. These results help to explain the general lack of a defined relationship between the abundance of C. raciborskii in freshwater ecosystems of Florida and observed concentrations of CYN. The latter observation raises the potential that previous reports of CYN may be coincidental with unrecorded presence of another CYN-producing species.     

The Palm Island mystery disease 20 years on: a review of research on the cyanotoxin cylindrospermopsin

Poisoning of humans resulting from consumption of water affected by the toxic cyanobacterium Cylindrospermopsis raciborskii was first reported almost 20 years ago from Palm Island, northern Queensland, Australia. Since that time a great deal has been learned about this organism and cylindrospermopsin (CYN), the toxin it produces. This article reviews the information now available to us. It summarizes aspects of the chemistry of the toxin-now known to be produced by some cyanobacterial species other than C. raciborskii-and its biosynthesis and chemical synthesis in vitro, as well as its detection and measurement by chemical and biological assay. Some of the factors affecting toxin production by cultured isolates of C. raciborskii are reviewed and the conditions that cause its release from the cells described. The occurrence of CYN in water bodies and the management strategies used to minimize the harmful effects of the toxin are outlined. These include a range of water-treatment practices now in place to remove CYN-producing organisms and/or to neutralize the toxin together with some management procedures that have been tried, with varying degrees of success, to prevent buildup of blooms of the offending organisms. Some of the public-health considerations arising from exposure to water supplies affected by CYN are summarized along with the risk factors and guidance values as they are currently applied. Among the more recent developments described are those that come from the application of molecular techniques for characterizing toxic and nontoxic strains and for exploring the genetic aspects of CYN production.     

Cylindrospermopsin occurrence in two German lakes and preliminary assessment of toxicity and toxin production of Cylindrospermopsis raciborskii (Cyanobacteria) isolates.

Cylindrospermopsis raciborskii, a freshwater cyanobacterium of tropical origin, is not only increasingly found in (sub) tropical water bodies, but also in temperate regions. Since this species may produce potent toxins such as cylindrospermopsin (CYN) and paralytic shellfish poisons, its massive occurrence in water bodies used as drinking water sources or for recreation is of major concern. The proliferation of C. raciborskii in German water bodies has been documented for the past decade. We investigated the occurrence of CYN in field populations and isolates of C. raciborskii from two lakes, and assessed the toxicity of culture isolates using the mouse bioassay, primary rat hepatocytes and human derived cell lines. We show for the first time the occurrence of CYN in German water bodies. None of seven isolates of C. raciborskii contained CYN, however, all isolates were toxic to primary rat hepatocytes, human hepatoblastoma (HEP-G2) and human colon adenocarcinoma (CACO-2) cells. Methanolic extracts were more toxic than aqueous extracts. Three isolates tested in the mouse bioassay were toxic at a concentration of 800 mg kg(-1) showing liver and spleen damage and inflammation of the intestine. These results give strong evidence that the German isolates of C. raciborskii contain currently not identified or unknown toxins.     

The accumulation of cylindrospermopsin from the cyanobacterium Cylindrospermopsis raciborskii in tissues of the Redclaw crayfish Cherax quadricarinatus.

Redclaw crayfish, Cherax quadricarinatus harvested from an aquaculture pond infested by a bloom of the cyanobacterium Cylindrospermopsis raciborskii (order: Nostocales), were shown to accumulate the toxic alkaloid cylindrospermopsin. Pond water samples collected during the bloom contained 589 microg l(-1) of the toxin (93% in the cyanobacterial cells, 7% in the water). Crayfish from the pond contained cylindrospermopsin at concentrations of 4.3 microg g freeze dried hepatopancreas tissue and 0.9 microg g freeze dried muscle tissue. Trichomes of C. raciborskii were observed in gut contents of crayfish harvested during the cyanobacterial bloom, indicating that the most likely mechanism for accumulation of the toxin was by ingestion of cyanobacterial cells. Crayfish subjected to an extract of harvested bloom material under laboratory conditions for a period of 14 days were also found to accumulate cylindrospermopsin, indicating that this toxin is also absorbed into the tissues by direct uptake of the toxin in solution.     

Toxicity of cylindrospermopsin to the brine shrimp Artemia salina: comparisons with protein synthesis inhibitors and microcystins.

The Artemia salina bioassay was successfully applied to the analysis of the hepatotoxic cyanobacterial alkaloid and protein synthesis inhibitor, cylindrospermopsin. A dose-dependent response in mortality was observed for purified cylindrospermopsin and LC(50) values decreased with time from 8.1 to 0.71 microg/ml(-1), between 24 and 72 h, respectively. Cylindrospermopsin was slightly less potent than micro cystin-LR, with similar LC(50) values on a gravimetric basis, but was more toxic to A.salina than the protein synthesis inhibitors, cycloheximide, chloramphenicol and tetracycline. Cylindrospermopsin-containing strains of the cyanobacterium Cylindrospermopsis raciborskii were found to be toxic to A.salina and the LC(50) concentration for these strains over time was greater than the LC(50) for purified cylindrospermopsin, with the exception of C. raciborskii strain CR1.     

Multiple-organ toxicity resulting from cylindrospermopsin exposure in tadpoles of the cane toad (Bufo marinus)

Histological examinations were made of cane toad (Bufo marinus) tadpoles after exposure to freeze-thawed Cylindrospermopsis raciborskii whole cell extracts and live C. raciborskii cultures containing sublethal concentrations of the blue-green algal toxin, cylindrospermopsin (CYN). Toxin exposure resulted in tissue injuries to multiple organs, with particular severity noted in the liver, intestine, nephric ducts and gill epithelia. The extent of cellular damage was similar across trials exposing tadpoles to aqueous and cell-bound toxins, despite unequal toxin concentrations being present in each. It was concluded that the presence of cell-bound toxin, which may be directly ingested via grazing, plays a crucial role in the exertion of histological effects in B. marinus. This work provides baseline information regarding the ecotoxicity of CYN toward amphibians. The range of cellular effects noted in CYN-exposed tadpoles suggests that toxic C. raciborskii blooms could represent considerable health risks to amphibian populations and indicate potentially far-reaching ecological impacts of toxic C. raciborskii blooms.     

Multiplex PCR assay for Cylindrospermopsis raciborskii and cylindrospermopsin-producing cyanobacteria

Water bodies are routinely monitored for the presence of potentially toxic cyanobacteria; however, the methodology for confirming toxicity is currently complex and expensive. Here we describe the application of gene-based technology to rapidly identify cylindrospermopsin-producing cyanobacteria, specifically, Cylindrospermopsis raciborskii. A multiplex polymerase chain reaction (PCR) test was developed that simultaneously identified polyketide synthase (pks) and peptide synthetase (ps) determinants associated with cylindrospermopsin production and distinguished C. raciborskii from other cylindrospermopsin-producing cyanobacteria of the species Anabaena bergii and Aphanizomenon ovalisporum, by targeting the rpoC1 gene. Twenty-one C. raciborskii, 5 A. bergii, 10 Aph. ovalisporum isolates and 3 environmental samples all yielded PCR results consistent with their toxicological status, as assessed by high-performance liquid chromatography coupled to mass spectrometry or matrix-assisted laser desorption ionization-time-of-flight mass spectrometry, and C. raciborskii was always correctly identified. The PCR test is a rapid, reliable, and economical way of assessing the toxic potential of cyanobacterial blooms formed by these organisms.     

Preliminary evidence for in vivo tumour initiation by oral administration of extracts of the blue-green alga cylindrospermopsis raciborskii containing the toxin cylindrospermopsin

New reports indicate that the toxic alkaloid cylindrospermopsin occurs in cyanobacteria in Israel, Florida, South America, and Australia in drinking water sources. This toxin is now recognised as a potential threat to human health. Furthermore, we have recently demonstrated the mutagenicity of cylindrospermopsin in vitro in a human lymphoblastoid cell-line. Therefore it is essential to determine whether cylindrospermopsin is also carcinogenic in vivo. In this preliminary study, 53 mice were treated up to three times orally with Cylindrospermopsis raciborskii extract containing cylindrospermopsin, while 27 control mice were treated with saline. A proportion of each group were then given O-tetradecanoylphorbol 13-acetate (10 microg/mouse, twice weekly in liquid food) for the duration of the experiment; the remainder were given a control diet. After 30 weeks, the mice were euthanased and the major organs were examined histologically. Five tumours were found in 53 cylindrospermopsin-treated mice while none were found in the 27 controls. Although the number of animals used was too low to provide statistical significance (p=0.16), the calculated relative risk (RR=6.2; 95% CI: 0.33-117) indicates a potential biological and public health significance requiring further investigation. Estimates are given of the size of experiment required to provide statistical proof of cylindrospermopsin carcinogenicity.     

Occurrence of the cyanobacterial toxin cylindrospermopsin in northeast Germany

The frequent occurrence of the cyanobacterial toxin cylindrospermopsin (CYN) in the (sub)tropics has been largely associated with cyanobacteria of the order Nostocales of tropical origin, in particular Cylindrospermopsis raciborskii. C. raciborskii is currently observed to spread northwards into temperate climatic zones. In addition, further cyanobacteria of the order Nostocales typically inhabiting water bodies in temperate regions are being identified as CYN-producers. Therefore, data on the distribution of CYN in temperate regions are necessary for a first assessment of potential risks due to CYN in water used for drinking and recreation. A total of 127 lakes situated in the north-eastern part of Germany were investigated in 2004 for the presence of the toxin CYN and the phytoplankton composition. The toxin could be detected in half of the lakes (n = 63) and in half of 165 samples (n = 88). Concentrations reached up to 73.2 microg CYN/g DW. CYN thus proved more widely distributed than previously demonstrated. The analyses of phytoplankton data suggest Aphanizomenon sp. and Anabaena sp. as important CYN producers in Germany, and confirm recent findings of Aphanizomenon flos-aquae as CYN-producing species frequently inhabiting water bodies in temperate climatic regions. The data shown here suggest that CYN may be an important cyanobacterial toxin in German water bodies and that further data are needed to assess this.     

Toxicity of the cyanobacterium Cylindrospermopsis raciborskii to Daphnia magna

The effect of two strains of Cylindrospermopsis raciborskii on the survivorship, somatic growth, and detoxification processes of juvenile Daphnia magna were investigated. Both strains of C. raciborskii (and also Ankistrodesmus falcatus, used as the control) were given to newborn D. magna at equivalent biovolumes. The survival curves for D. magna subjected to the two C. raciborskii treatments differed from those of the starved and fed treatments. After 48 h of exposure, the percentage of D. magna surviving after exposure to Cylin-A (a cylindrospermopsin-producing strain isolated from Australia) and Cylin-P (a non-cylindrospermopsin-producing strain isolated from Portugal) was 10.00% and 93.33%, respectively. The strain that produces cylindrospermopsin caused the greatest toxic effect in juvenile D. magna. Statistically significant differences in D. magna body size between the four treatments (Cylin-A, Cylin-P, A. falcatus, and starved) were detected after 48 h of exposure. The juvenile D. magna that received the two C. raciborskii treatments showed an increase in size (relative to their size at T(0)) of 2.54% and 38.14%, respectively. These values were statistically significantly different than those of the A. falcatus-fed control (55.54%) and the starved control (11.47%). In both C. raciborskii treatments there was a tendency for increased GST enzyme activities after 24 h of exposure. Cylindrospermopsin was detected (HPLC-MS/MS) in D. magna tissues after 24 and 48 h (0.025 and 0.02 ng animal(-)1, respectively). The results of this study indicate that C. raciborskii can affect the fitness and growth potential of juvenile D. magna.     

锡林郭勒盟高原盐湖放线菌资源勘探及抗菌活性筛选

摘要：目的 探究锡林郭勒盟高原盐湖放线菌的多样性、新颖性及抗菌活性,为新型抗菌活性产物的发掘储备菌种资源。方 法 采用20种分离培养基,以平板涂布法分离放线菌;依据16S rRNA基因的同源性对菌株进行分子鉴定;PCR检测代表菌株的I型聚 酮合酶(PKS I)、II型聚酮合酶(PKS II)和非核糖体多肽合成酶(NRPS)功能基因;菌株的发酵液上清和菌体分别经乙酸乙酯和丙酮提取, 提取样品经纸片扩散法进行抗菌活性检测。目标菌株合成次级代谢产物的能力通过antiSMASH对基因组序列进行分析预测。结果 从 7份盐湖土壤样品中分离获得了365株放线菌,其分布于放线菌纲的8个目15个科28个属,优势菌属为链霉菌属和拟诺卡菌属。分离获 得的链霉菌新颖性突出,13株链霉菌与近缘菌株16S rRNA基因的最高相似度≤98.0%,推测其归属于10个链霉菌属新种。受试放线菌 对革兰阳性菌的抗菌活性显著,并从中筛选出3株抗菌作用强且抗菌谱较广的链霉菌;20株受试放线菌同时含有3种抗生素生物合成基 因。菌株XMNu-295基因组含有24个潜在的次级代谢产物生物合成基因簇,以聚酮类、非核糖体多肽类和萜烯类等为主。结论 锡林 郭勒盟高原盐湖中可培养放线菌多样性较为丰富,新颖性突出,目标菌株值得进一步开展次级代谢产物的化学研究。     

Influence of intracellular toxin concentrations on cylindrospermopsin bioaccumulation in a freshwater gastropod (Melanoides tuberculata).

Scant information is available regarding the bioaccumulation of cylindrospermopsin (CYN) in aquatic organisms, particularly in invertebrates. This study examined toxin bioconcentration and bioaccumulation in the aquatic snail, Melanoides tuberculata, following exposure to freeze-thawed whole cell extracts and a live Cylindrospermopsis raciborskii culture containing CYN. Both bioconcentration and bioaccumulation were evident, but exposure to toxin in the freeze-thawed solutions resulted in minor tissue contamination compared with that resulting from live C. raciborskii exposure. Thus, whilst CYN uptake resulted from both extracellular and intracellular exposures, the availability of intracellular toxin was critical in affecting tissue CYN values. M. tuberculata did not bioconcentrate CYN into the shell. Bioaccumulation of the analog deoxy-CYN was also recorded. Knowledge of intracellular toxin concentrations may be critical in evaluating the bioaccumulation, ecological and human health risks associated with contaminated systems.     

Degradation of the cyanobacterial toxin cylindrospermopsin, from Cylindrospermopsis raciborskii, by chlorination.

Cylindrospermopsin, a potent cyanobacterial toxin produced by Cylindrospermopsis raciborskii and other cyanobacteria, is regularly found in water supplies of Queensland, Australia. This study focussed on the effectiveness of chlorination as a water treatment procedure for cylindrospermopsin degradation. The results demonstrate that relatively low chlorine doses (<1 mg l(-1)) are sufficient for degradation of cylindrospermopsin, when the dissolved organic carbon content is low. However, if organic matter other than cylindrospermopsin is present in the solution, the effectiveness of chlorine for cylindrospermopsin degradation is reduced as other organic matter present consumes chlorine. Under the experimental conditions using samples with a solution pH of 6-9, a residual chlorine concentration of 0.5 mg l(-1)99% of cylindrospermopsin. Toxin degradation via chlorination occurs within the first minute and no difference was observable between degradation in an open system and in a closed system. With a decrease of the pH from 6 to 4 a reduction in the efficiency of chlorine for degradation of cylindrospermopsin was observable, a possible indication that cylindrospermopsin is more stable to chlorine degradation at lower pH. However, in normal water treatment this is not relevant since the pH is consistently higher than 6.     

Cyanobacterial (blue-green algal) toxins in water supplies: Cylindrospermopsins

The toxic alkaloid cylindrospermopsin is produced by a range of cyanobacterial species worldwide. It was first identified in the species Cylindrospermopsis raciborskii from tropical waters, and has since been isolated from four other genera in locations ranging from Israel to Japan. High concentrations of the organisms and toxin have been identified in reservoirs, natural lakes, and rivers in summer in the USA and in Australia. The toxin is a particular problem in drinking water sources as concentrations in the free water are appreciable, so that removal of the filaments during water treatment does not remove the toxin. The toxicity resulting from oral ingestion is seen in the liver, kidneys, stomach, intestine, and white blood cells, with some vascular damage in mice. Gastrointestinal as well as liver injury has been observed in human poisoning. Studies of toxicity in vitro have shown inhibition of protein synthesis. Genotoxicity has also been demonstrated, and there is preliminary evidence for carcinogenicity. A Guideline Value for safe water supply of 1 microg/L has been proposed. Research into toxin measurement techniques and water treatment methods has indicated that effective control measures may be practicable for this toxin in drinking water. Considerably more research is needed to fully define the health risks from this toxin.     

Cylindrospermopsin, a cyanobacterial alkaloid: evaluation of its toxicologic activity

This paper describes the natural occurrence of the toxin, cylindrospermopsin, in two species of cyanobacteria found in Australia. The structure and chemical properties of this compound are described along with a nontoxic analog of cylindrospermopsin. The results of both intraperitoneal (IP) and oral dosing of mice show that hepatotoxicity is the main effect of cylindrospermopsin in vivo, but that a thrombohemorrhagic phenomenon is observed in a proportion of dosed animals. It has been shown that the toxin can be metabolized in vivo and that a bound metabolite occurs in the liver. Cytotoxicity experiments using cell cultures show that cylindrospermopsin is more cytotoxic to isolated rat liver hepatocytes than to other cell types. Risk assessment calculations show that guideline values for cylindrospernopsin in drinking water should lie in the low microgram per liter range.     

Effects of Cylindrospermopsis raciborskii and Aphanizomenon ovalisporum (cyanobacteria) ingestion on Daphnia magna midgut and associated diverticula epithelium

This article reports a light and electron microscopy investigation of the effects of Cylindrospermopsis raciborskii and Aphanizomenon ovalisporum ingestion on midgut and associated digestive diverticula of Daphnia magna. Additionally, survivorship and growth effects caused by feeding on cyanobacteria were assessed. Three cyanobacteria were used in the experiments: cylindrospermopsin (CYN)-producing C. raciborskii, CYN-producing A. ovalisporum and non-CYN-producing C. raciborskii. In order to discriminate between the alterations due to the low nutritional value of cyanobacteria and toxic effects, a control group was fed on the chlorophyte Ankistrodesmus falcatus and another control group was not fed. In the chlorophyte fed control, the epithelium lining the midgut and associated diverticula is mainly formed by strongly stained cells with an apical microvilli border. Nevertheless, unstained areas in which cell lyses had occurred were also observed. In the unfed control, the unstained areas became predominant due to an increment of cell lyses. All individuals fed on CYN-producing A. ovalisporum and some of those fed on non-CYN-producing C. raciborskii appear similar to the unfed control. However, some individuals fed on non-CYN-producing C. raciborskii showed similarities with the fed control. In contrast, the midgut and digestive diverticula of D. magna fed on CYN-producing C. raciborskii showed a widespread dissociation of epithelial cells, associated with severe intracellular disorganization, but cell lysis was less evident than in controls. These alterations cannot be attributed to CYN, because those effects were not induced by CYN-producing A. ovalisporum. Therefore, data suggest the production of another unidentified active metabolite by CYN-producing C. raciborskii, responsible for the disruption of cell adhesion in the epithelium of D. magna digestive tract. Data also show that the tested cyanobacteria are inadequate as food to D. magna, due to low nutritional value and toxic content.     

First report on cylindrospermopsin producing Aphanizomenon flos-aquae (Cyanobacteria) isolated from two German lakes.

Three single-filament isolates of Aphanizomenon flos-aquae from two German lakes were found to produce remarkable amounts of the cyanobacterial hepatotoxin cylindrospermopsin (CYN). CYN-synthesis of the strains were evidenced both by LC-MS/MS analysis and detection of PCR products of gene fragments which are implicated in the biosynthesis of the toxin. The strains contain CYN in the range of 2.3-6.6 mg g(-1) of cellular dry weight. To our knowledge this is the first report of CYN in A. flos-aquae.     

Studies of the comparative in vitro toxicology of the cyanobacterial metabolite deoxycylindrospermopsin

Cyanobacteria are capable of producing metabolites that are in some cases toxic to humans and other animals. Of these metabolites, the toxin cylindrospermopsin (CYN) is produced by a number of species of cyanobacteria including Cylindrospermopsis raciborskii, and its toxicity has been documented. The CYN analog deoxycylindrospermopsin (deoxyCYN) is commonly produced in varying proportions by the cyanobacteria that produce CYN. The toxicological profile of CYN suggests that it is primarily a hepatotoxin, but with the capacity to damage other organs and tissues. Limited in vivo information is available on the toxicity of deoxyCYN and suggests it to be of low potency. The aim of this research was to determine the comparative toxicology of deoxyCYN using in vitro systems. Using cell viability assays, it was shown that deoxyCYN had inhibitory effects on cell viability and proliferation of a similar magnitude to that of CYN. Morphological changes in deoxyCYN-treated cells were similar to those of CYN. Investigation of protein synthesis inhibition demonstrated that deoxyCYN was of similar potency to CYN. Inhibition of protein synthesis is an acknowledged mechanism of toxicity for CYN, and the results produced here suggest that deoxyCYN operates by similar toxicological mechanisms to CYN and that in vivo animal testing should be undertaken to clarify the potential for risk to humans from this toxin.     

Cell-free protein synthesis inhibition assay for the cyanobacterial toxin cylindrospermopsin

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% confidence limits (Cl) = 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 microM (200-1200 micrograms/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.