Effects and possible mechanisms of sanguinarine on the competition between Raphidiopsis raciborskii (Cyanophyta) and Scenedesmus obliquus (Chlorophyta): A comparative toxicological study.

The phytogenic algicide sanguinarine shows strong inhibitory effects on some bloom-forming cyanobacteria and exhibits great potential in cyanobacterial bloom mitigation. To evaluate the possible ecological effects of sanguinarine on microalgae, the effects and possible mechanisms of sanguinarine on the competition between bloom-forming cyanobacterium Raphidiopsis raciborskii (formerly named Cylindrospermopsis raciborskii) and green alga Scenedesmus obliquus were investigated through co-culture competition test and comparative toxicological study including growth characteristics, chlorophyll fluorescence transients, activities of antioxidant enzymes, and lipid peroxidation. The results of Raphidiopsis-Scenedesmus co-culture competition test showed that sanguinarine decreased the competition ability of R. raciborskii, which benefitted S. obliquus in winning the competition. Toxicological studies have shown that sanguinarine exhibited high inhibitory effects on the growth and photosynthesis of R. raciborskii but no obvious toxicity on S. obliquus at concentrations of no more than 80 µg L-1. Oxidative damage partially contributed but was not the primary mechanism for the toxicity of sanguinarine on R. raciborskii. The results presented in this study indicate that sanguinarine may be a good algicidal candidate in mitigation of Raphidiopsis-based water bloom.

Clethodim (herbicide) alters the growth and toxins content of Microcystis aeruginosa and Raphidiopsis raciborskii.

Increased agricultural intensification goes with the widespread use of herbicides that adversely affect aquatic biodiversity. The effects of herbicides on toxin-producing cyanobacteria have been poorly studied. The present study aimed to investigate the toxicological and physiological effects of the herbicide clethodim on Raphidiopsis raciborskii (a.k.a. Cylindrospermopsis raciborskii) ITEPA1 and Microcystis aeruginosa BCCUSP232. On day four of the experiment, the exposure to 25 mg/L clethodim resulted in the highest cell density of R. raciborskii. Similarly, exposure to the 1, 5, 20, and 50 mg/L clethodim treatments resulted in the highest cell densities of M. aeruginosa on day 4 of the experiment. Medium effect concentrations (EC50) after 96 h of exposure of both strains to clethodim were 192.98 mg/L and 168.73 mg/L for R. raciborskii and M. aeruginosa, respectively. The presence of clethodim significantly increased the total microcystin content of M. aeruginosa compared to the control cultures. At 400 mg/L, total saxitoxins content of R. raciborskii was 27% higher than that of the control cultures on day 4. In contrast, cultures exposed to 100 mg/L clethodim had the lowest saxitoxins levels per cell quota. There was an increase in the levels of intracellular hydrogen peroxide in both species during exposure to clethodim, which was followed by significant changes (p < 0.05) in the activity of antioxidant enzymes such as peroxidase and superoxide dismutase. These results revealed that the presence of low levels of clethodim in the aquatic environment might lead to the excessive proliferation of cyanobacteria and alteration of their cyanotoxins content.

Physiological responses of the freshwater N2 -fixing cyanobacterium Raphidiopsis raciborskii to Fe and N availabilities.

The cyanobacterium Raphidiopsis raciborskii is of environmental and social concern in view of its toxicity, bloom-forming characteristics and increasingly widespread occurrence. However, while availability of macronutrients and micronutrients such as N and Fe are critically important for the growth and metabolism of this organism, the physiological response of toxic and non-toxic strains of R. raciborskii to varying Fe and N availabilities remains unclear. By determining physiological parameters as a function of Fe and N availability, we demonstrate that R. raciborskii growth and N2 -fixing activity are facilitated at higher Fe availability under N2 -limited conditions with faster growth of the CS-506 (cylindrospermopsin-producing) strain compared with that of CS-509 (the non-toxic) strain. Radiolabelled Fe uptake assays indicated that R. raciborskii acclimated under Fe-limited conditions acquires Fe at significantly higher rates than under Fe replete conditions, principally via unchelated Fe(II) generated as a result of photoreduction of complexed Fe(III). While N2 -fixation of both strains occurred during both day and night, the CS-506 strain overall exhibited higher N2 -fixing and Fe uptake rates than the CS-509 strain under N-deficient and Fe-limited conditions. The findings of this study highlight that Fe availability is of significance for the ecological advantage of CS-506 over CS-509 in N-deficient freshwaters.

Chlorophyll a fluorescence and transcriptome reveal the toxicological effects of bisphenol A on an invasive cyanobacterium, Cylindrospermopsis raciborskii.

Bisphenol A has attracted worldwide attention due to its harmful effects on humans, animals and plants. In this study, the toxicological effects of BPA on Cylindrospermopsis raciborskii were assessed based on chlorophyll a fluorescence and transcriptome analyses. The results showed that the growth of C. raciborskii was significantly inhibited when BPA exceeded 0.1 mg L-1. A marked rise of phase J was observed at a concentration greater than 0.1 mg L-1, while a K phase appeared at 20 mg L-1. The chlorophyll a fluorescence parameters of RC/CS0, F0, φP0, φE0, and ψ0, underwent a significant decline under all treatments of BPA, whereas a significant increase in both VJ and M0 occurred under all concentrations of BPA. Additionally, ABS/RC and DIo/RC markedly increased at 10 mg L-1 and 20 mg L-1. The transcriptome analysis revealed that the genes of photosynthesis, including psbA, psbB, psbC, psbD, apcA, apcB, cpcA, and cpcB, as well as those of chlorophyll and carotenoid biosynthesis, namely hemN, acsF, chlL, chlN, chlP, crtB, pds, were all down-regulated. Moreover, BPA also inhibited the oxidative phosphorylation, glycolysis/gluconeogenesis, citrate cycle (TCA cycle), and fatty acid metabolism in C. raciborskii. Taken together, these results suggest BPA can negatively affect the expression of multiple genes and the vital energy metabolism process to arrest the growth and photosynthesis of C. raciborskii.

The Effect of Bisphenol A on Growth, Morphology, Lipid Peroxidation, Antioxidant Enzyme Activity, and PS II in Cylindrospermopsis raciborskii and Scenedesmus quadricauda.

To investigate the effect of bisphenol A (BPA) on Cylindrospermopsis raciborskii (Cyanobacteria) and Scenedesmus quadricauda (Chlorophyta), we grew the two species at BPA concentrations of 0, 0.1, 1, 2, 5, 10, and 20 mg/L and examined their growth, lipid peroxidation, antioxidant enzyme activity, and chlorophyll a fluorescence. The 96-h EC50 values (effective concentration causing 50% growth inhibition) for BPA in C. raciborskii and S. quadricauda were 9.663 ± 0.047, and 13.233 ± 0.069 mg/L, respectively. A significant reduction in chlorophyll a concentration was found in C. raciborskii and S. quadricauda when BPA concentrations were greater than 1 and 2 mg/L, respectively. Furthermore, F v/F m, ΔF/F m', and qP decreased significantly at 10 mg/L BPA in C. raciborskii but started to decrease at 10 mg/L in S. quadricauda. The changes in chlorophyll fluorescence parameters (α, rETRmax) that were obtained from the rapid light response curves of both algae species showed similar responses to F v/F m, ΔF/F m', and qP under BPA-induced stress. Values for all of the chlorophyll fluorescence parameters in S. quadricauda were higher than in C. raciborskii; however, the nonphotochemical quenching measured in C. raciborskii was considerably higher than it was in S. quadricauda. In addition, lipid peroxidation (determined as MDA content) and antioxidant enzyme activities (SOD and CAT) increased in both species as the BPA concentration increased. These results suggest that C. raciborskii is more sensitive to the effects of BPA than S. quadricauda and that photosystem II might be a target for the activity of BPA in vivo.

The efficiency of combined coagulant and ballast to remove harmful cyanobacterial blooms in a tropical shallow system.

We tested the hypothesis that a combination of coagulant and ballast could be efficient for removal of positively buoyant harmful cyanobacteria in shallow tropical waterbodies, and will not promote the release of cyanotoxins. This laboratory study examined the efficacy of coagulants [polyaluminium chloride (PAC) and chitosan (made of shrimp shells)] alone, and combined with ballast (lanthanum modified bentonite, red soil or gravel) to remove the natural populations of cyanobacteria collected from a shallow eutrophic urban reservoir with alternating blooms of Cylindrospermopsis and Microcystis. PAC combined with ballast was effective in settling blooms dominated by Microcystis or Cylindrospermopsis. Contrary to our expectation, chitosan combined with ballast was only effective in settling Cylindrospermopsis-dominated blooms at low pH, whereas at pH≥8 no effective flocculation and settling could be evoked. Chitosan also had a detrimental effect on Cylindrospermopsis causing the release of saxitoxins. In contrast, no detrimental effect on Microcystis was observed and all coagulant-ballast treatments were effective in not only settling the Microcystis dominated bloom, but also lowering dissolved microcystin concentrations. Our data show that the best procedure for biomass reduction also depends on the dominant species.

Pharmacological studies confirm neurotoxic metabolite(s) produced by the bloom-forming Cylindrospermopsis raciborskii in Hungary.

A rapid cyanobacterial bloom of Cylindrospermopsis raciborskii (3.2 × 10(4) filaments/mL) was detected early November, 2012, in the Fancsika pond (East Hungary). The strong discoloration of water was accompanied by a substantial fish mortality (even dead cats were seen on the site), raising the possibility of some toxic metabolites in the water produced by the bloom-forming cyanobacteria (C. raciborskii). The potential neuronal targets of the toxic substances in the bloom sample were studied on identified neurons (RPas) in the central nervous system of Helix pomatia. The effects of the crude aqueous extracts of the Fancsika bloom sample (FBS) and the laboratory isolate of C. raciborskii from the pond (FLI) were compared with reference samples: C. raciborskii ACT 9505 (isolated in 1995 from Lake Balaton, Hungary), the cylindrospermopsin producer AQS, and the neurotoxin (anatoxin-a, homoanatoxin-a) producer Oscillatoria sp. (PCC 6506) strains. Electrophysiological tests showed that both FBS and FLI samples as well the ACT 9505 extracts modulate the acetylcholine receptors (AChRs) of the neurons, evoking ACh agonist effects, then inhibiting the ACh-evoked neuronal responses. Dose-response data suggested about the same range of toxicity of FBS and FLI samples (EC50 = 0.397 mg/mL and 0.917 mg/mL, respectively) and ACT 9505 extracts (EC50 = 0.734 mg/mL). The extract of the neurotoxin-producing PCC 6506 strain, however, proved to be the strongest inhibitor of the ACh responses on the same neurons (EC50 = 0.073 mg/mL). The presented results demonstrated an anatoxin-a-like cholinergic inhibitory effects of cyanobacterial extracts (both the environmental FBS sample, and the laboratory isolate, FLI) by some (yet unidentified) toxic components in the matrix of secondary metabolites. Previous pharmacological studies of cyanobacterial samples collected in other locations (Balaton, West Hungary) resulted in similar conclusions; therefore, we cannot exclude that this chemotype of C. raciborskii which produce anatoxin-a like neuroactive substances is more widely distributed in this region.

Elevated CO2 causes changes in the photosynthetic apparatus of a toxic cyanobacterium, Cylindrospermopsis raciborskii.

We studied the physiological acclimation of growth, photosynthesis and CO2-concentrating mechanism (CCM) in Cylindrospermopsis raciborskii exposed to low (present day; L-CO2) and high (1300ppm; H-CO2) pCO2. Results showed that under H-CO2 the cell specific division rate (µc) was higher and the CO2- and light-saturated photosynthetic rates (Vmax and Pmax) doubled. The cells' photosynthetic affinity for CO2 (K0.5CO2) was halved compared to L-CO2 cultures. However, no significant differences were found in dark respiration rates (Rd), pigment composition and light harvesting efficiency (α). In H-CO2 cells, non-photochemical quenching (NPQ), associated with state transitions of the electron transport chain (ETC), was negligible. Simultaneously, a reorganisation of PSII features including antenna connectivity (JconPSIIα), heterogeneity (PSIIα/ß) and effective absorption cross sectional area (σPSIIα/ß) was observed. In relation to different activities of the CCM, our findings suggest that for cells grown under H-CO2: (1) there is down-regulation of CCM activity; (2) the ability of cells to use the harvested light energy is altered; (3) the occurrence of state transitions is likely to be associated with changes of electron flow (cyclic vs linear) through the ETC; (4) changes in PSII characteristics are important in regulating state transitions.

Cylindrospermopsis raciborskii dominates under very low and high nitrogen-to-phosphorus ratios.

In freshwater ecosystems, a variety of factors mediate phytoplankton community structure, including herbivore community structure, light availability, temperature, mixing, and absolute and relative nutrient concentrations (total nitrogen (TN), total phosphorus (TP)). Ecological stoichiometry examines how the nutrient content of organisms and their environment may mediate population-, community-, and ecosystem-level processes. The manipulation of N:P ratios is a widely regarded tool for managing phytoplankton species composition given that nitrogen-fixing cyanobacteria should dominate algal communities under relatively low N:P (<64:1, by atoms) given their ability to convert dissolved dinitrogen gas into organic nitrogen. However, due to the physiological expense of nitrogen fixation, diazotrophs should be outcompeted by non-nitrogen fixing phytoplankton under higher N:P when other environmental factors are similar. We tested this hypothesis in a field experiment using 2500-L limnocorrals installed in a eutrophic lake (ambient N:P ∼40:1 (by atoms); TN ∼1360 µgL(-1); TP ∼75 µgL(-1)). At the start of the experiment, we randomly assigned limnocorrals among the ambient (40:1) and low (7:1) or high (122:1) N:P treatments (n = 4 replicates/treatment), which were established by adding P or N at the start of the experiment, respectively. The phytoplankton community in the enclosures at the start of the experiment was diverse (i.e., 18 phytoplankton genera) and dominated by chlorophytes (including Coelastrum and Scenedesmus (30% and 13% of total biomass, respectively)) and cyanobacteria (including Anabaena and Cylindrospermopsis (23% and 17% of total biomass, respectively)). In contrast to predictions based on ecological stoichiometry, the phytoplankton community in all N:P treatments increased in abundance and was almost entirely composed of the nitrogen-fixing cyanobacterium, Cylindrospermopsis raciborskii, by the conclusion of the study. Moreover, concentrations of the cyanobacterial neurotoxin, saxitoxin, were enhanced under the two highest N:P conditions. The ability of C. raciborskii to dominate phytoplankton communities under such extreme N:P shows that short-term management of nutrient stoichiometry through fertilization is not likely to be effective for controlling blooms of this noxious cyanobacterium and may help to explain the rapid expansion of this invasive species to temperate latitudes.

Cylindrospermopsis raciborskii exudate-Cu complexes: impact on copper dynamics and bioavailability in an aquatic food chain.

INTRODUCTION: The increasing contamination of aquatic environments motivates studies on the interactions among natural dissolved organic matter, metals, and the biota. This investigation focused on the organic exudates of the toxic cyanobacteria Cylindrospermopsis raciborskii as a Cu carrier through a three-level aquatic trophic chain (bacteria, protozoa, and copepod). DISCUSSION: The effects of bacteria activity and growth on the metal-organic complexes were evaluated through changes in free Cu(2+) ions, total dissolved, and total particulate Cu. To be sure that the added copper would be complexed to the exudates, its complexing properties were previously determined. The cyanobacteria exudate-Cu complexes were furnished to bacteria that were further used as a food source to the protozoan Paramercium caudatum. This was then furnished as food to the copepod Mesocyclops sp. The results showed that, in general, the cyanobacterial exudates decreased Cu bioavailability and toxicity to the first trophic level (bacteria), but because the heterotrophic bacteria accumulated Cu, they were responsible for the transference for the otherwise low availability metal form. Both the bacteria and protozoan organisms accumulated Cu, but no metal accumulation was detected in the copepods.

The impacts of a high CO2 environment on a bicarbonate user: the cyanobacterium Cylindrospermopsis raciborskii.

The potentially toxic cyanobacterium Cylindrospermopsis raciborskii (Woloszynska) Seenya et Subba Raju, originally described as a tropical-subtropical species, is increasingly found in temperate regions and its range is expanding. Climate change is hypothesised to be a factor in this expansion. We studied the effects of dissolved inorganic carbon (DIC) and pH on growth and photosynthesis of this species. We prepared six treatments in a continuous culture (turbidostat) grown at high light, two at low light, and eight in batch cultures grown under low light, by manipulating pH, HCO(3)(-) and CO(2) to assess the effect, if any, of these parameters on the growth rate, inorganic carbon acquisition and photosynthetic parameters of C. raciborskii. When the turbidostats were grown at 100 µmol photons (PAR) m(-2) s(-1), HCO(3)(-) concentration and pH had a positive effect on growth rate; the specific growth rate in 6 mM HCO(3)(-), for example, was twice what it was in 0.6 mM HCO(3)(-) (0.84 ± 0.10 and 0.44 ± 0.04 d(-1) respectively). Growth was lower in turbidostats grown at 20 µmol photons (PAR) m(-2) s(-1). Isotope disequilibrium experiments showed that the contribution of HCO(3)(-) to DIC acquisition is proportionately greater at the higher light. The maximum relative electron transport rate (rETR(max)) was significantly higher at the higher light, while the slope of the linear portion of the rETR(max) versus irradiance curve (α) was unchanged. In low light batch cultures, increasing HCO(3)(-) concentration and pH had a negative effect on growth, while CO(2) concentration had a small, positive effect. This species of cyanobacterium has an efficient CCM and under ideal growing conditions gets most of its carbon from HCO(3)(-). It may, therefore, be at a competitive disadvantage in a high CO(2) world.