Phosphorus limitation combined with aluminum triggers synergistic responses on the freshwater microalgae Raphidocelis subcapitata (Chlorophyceae).

In the environment, algae are exposed to several stressors such as limitation of essential nutrients and excess of toxic substances. It is well known the importance of phosphorus (P) supply for healthy metabolism of algae and impacts at this level can affect the whole aquatic trophic chain. Aluminum (Al) is the most abundant metal on Earth and it is toxic to different trophic levels. Processes related to P and Al assimilation still need to be clarified and little is known about the responses of microalgae exposed to the two stressors simultaneously. We evaluated the effects of environmental concentrations of Al and P limitation, isolated and in combination, on growth, pigment production and photosynthesis of the freshwater microalga Raphidocelis subcapitata. Both stressors affected cell density, chlorophyll a, carotenoids, and maximum quantum yield. Al did not affect any other evaluated parameter, while P limitation affected parameters related to the dissipation of heat by algae and the maximum electron transport rate, decreasing the saturation irradiance. In the combination of both stressors, all parameters evaluated were affected in a synergistic way, i.e., the results were more harmful than expected considering the responses to isolated stressors. Our results indicate that photoprotection mechanisms of algae were efficient in the presence of both stressors, avoiding damages to the photosynthetic apparatus. In addition, our data highlight the higher susceptibility of R. subcapitata to Al in P-limited conditions.

Effect of phosphorus on the toxicity of zinc to the microalga Raphidocelis subcapitata.

The aim of this study was to evaluate the effect of phosphorus (P) on the toxicity of zinc (Zn) for the alga Raphidocelis subcapitata. P was provided in three concentrations: 2.3 x 10-4 mol L-1, 2.3 x 10-6 mol L-1 and 1.0 x 10-6 mol L-1. Algal cells were acclimated to the specific P concentrations before the start of the experiment. The chemical equilibrium software MINEQL+ 4.61 was employed to calculate the Zn2+ concentration. After acclimated, the algal cells were inoculated into media containing different Zn concentrations (0.09 x 10-6 mol L-1 to 9.08 x 10-6 mol L-1). The study showed that besides the reduction in algal growth rates, phosphorus had an important influence on the toxicity of zinc for microalga. The inhibitory Zn2+ concentration values for R. subcapitata were 2.74 x 10-6 mol L-1, 0.58 x 10-6 mol L-1 and 0.24 x 10-6 mol L-1 for the microalgae acclimated at P concentrations of 2.3 x 10-4 mol L-1, 2.3 x 10-6 mol L-1 and 1.0 x 10-6 mol L-1, respectively. Ecotoxicological studies should consider the interaction between metal concentrations and varying P values to provide realistic data of what occurs in phytoplankton communities in environments.

Potential effects of fungicide and algaecide extracts of Annona glabra L. (Annonaceae) on the microalgae Raphidocelis subcapitata and on the oomycete Pythium.

Annona glabra L. is a semi-deciduous tree that contains several active substances, including secondary metabolites, with antifungal activity. Phytopathogenic strains of the genus Pythium cause billion dollar losses all over the world on natural and crop species. Searching for eco-friendly algaecides and fungicides, we analyzed the effects of acetone extracts of A. glabra leaves on the algae Rhaphidocelis subcapitata (Korshikov) and on the oomycete Pythium aphanidermatum (Edson). We evaluated ten extract concentrations for each organism - 0 to 400 mg L-1 for algae and 0-1000 µg disc1 for oomycete. The results showed no effect on algae up to 75 mg L-1, but a significant inhibitory effect at 125 mg L-1 and above, which reduced the growth rate and the final biomass of the algae. Extract concentrations above 200 mg L-1 were completely inhibitory. The half maximal inhibitory concentration for 72 and 96 h of exposure to our crude extracts are comparable to those obtained with commercial fungicides and herbicides used in aquatic ecosystems. The P. aphanidermatum inhibition concentrations have effects comparable to fungicides as Cycloheximide and Bifonazole. Some substances isolated from the extracts are described as antifungals, which could explain part of anti-oomycete activity. Our results highlight the importance of searching bioactive compounds from plants.

Potential effects of fungicide and algaecide extracts of Annona glabra L. (Annonaceae) on the microalgae Raphidocelis subcapitata and on the oomycete Pythium

ABSTRACT Annona glabra L. is a semi-deciduous tree that contains several active substances, including secondary metabolites, with antifungal activity. Phytopathogenic strains of the genus Pythium cause billion dollar losses all over the world on natural and crop species. Searching for eco-friendly algaecides and fungicides, we analyzed the effects of acetone extracts of A. glabra leaves on the algae Rhaphidocelis subcapitata (Korshikov) and on the oomycete Pythium aphanidermatum (Edson). We evaluated ten extract concentrations for each organism - 0 to 400 mg L-1 for algae and 0-1000 µg disc1 for oomycete. The results showed no effect on algae up to 75 mg L-1, but a significant inhibitory effect at 125 mg L-1 and above, which reduced the growth rate and the final biomass of the algae. Extract concentrations above 200 mg L-1 were completely inhibitory. The half maximal inhibitory concentration for 72 and 96 h of exposure to our crude extracts are comparable to those obtained with commercial fungicides and herbicides used in aquatic ecosystems. The P. aphanidermatum inhibition concentrations have effects comparable to fungicides as Cycloheximide and Bifonazole. Some substances isolated from the extracts are described as antifungals, which could explain part of anti-oomycete activity. Our results highlight the importance of searching bioactive compounds from plants.

Copper affects biochemical and physiological responses of Selenastrum gracile (Reinsch).

Copper is an essential metal for several physiological and metabolic processes, but a narrow range regulate its effect in phytoplankton cells. It can affect the production of biomolecules and be toxic at concentrations slightly above those required, e.g. decreasing photosynthesis and increasing respiration. The aims of this study were to analyse the changes in growth and chlorophyll a synthesis, and in biochemistry (total carbohydrates, proteins, lipids and fatty acids) of the freshwater microalga Selenastrum gracile after exposure to copper. Exponentially growing cells were exposed to 5 concentrations of free copper ions (Cu2+) ranging from 0.7 (control) to 13 × 10-8 M for up to 120 h. Free Cu2+ ion concentrations were calculated through the chemical equilibrium model MINEQL+. We observed that copper was responsible for a decrease in cell density and an increase in total protein and lipid production, but no effect on total carbohydrates was detected. The increase in phospholipids and sterols and a decrease in saturated fatty acids under copper exposure suggest a change in conformation of the cell membrane, by decreasing its fluidity. We suggest this serves the cell as a system to avoid the internalization of metal, thereby acting as a detoxifying mechanism.

Effect of copper contaminated food on the life cycle and secondary production of Daphnia laevis.

In aquatic environments, copper (Cu) plays important physiological roles in planktonic food chain, such as electron transfer in photosynthesis and constituting proteins that transport oxygen in some arthropods, while at higher concentrations it is toxic on these organisms and higher trophic levels. The combined effects of natural (e.g. volcanic activity) and anthropogenic sources (e.g. mining waste) contribute to the increase in copper pollution in different ecosystems and regions around the world. In the present study, we evaluated the bioaccumulation and effect of Cu on Raphidocelis subcapitata (freshwater algae), and the influence of Cu-contaminated food (algae) on Daphnia laevis (tropical cladoceran). The amount of copper accumulated in microalgae and cladoceran was quantified, and life-history parameters of D. laevis such as growth, reproduction and longevity were measured. The cell density of Cu exposed R. subcapitata declined, and cladoceran fed with contaminated food had lower longevity, production of eggs and neonates, and reduced secondary production. A concentration dependent increase in Cu accumulation was observed in the microalgae, while the opposite occurred in the animal, indicating a cellular metal regulatory mechanism in the latter. However, this regulation seems not to be sufficient to avoid metal induced damages in the cladoceran such as decreased longevity and reproduction. We conclude that diet is an important metal exposure route to this cladoceran, and the assessment of chronic contamination during the complete life cycle of cladoceran provides results that are similar to those observed in natural environments, especially when native organisms are investigated.

Influence of phosphorus on copper toxicity to Selenastrum gracile (Reinsch) Korshikov.

Microalgae need a variety of nutrients for optimal growth and health. However, this rarely occurs in nature, and if nutrient proportions vary, biochemical changes can occur in phytoplankton community. This may result in modifications of zooplankton food quality, affecting aquatic food chains. Our aim was to investigate the toxicity of copper (Cu) to Selenastrum gracile, a common freshwater Chlorophyceae, at different physiological status induced by varying phosphorus (P) concentration in culture medium. Phosphorus was investigated at 2.3×10(-4), 1.1×10(-4), 2.3×10(-5), 4.6×10(-6) and 2.3×10(-6) mol L(-1) and Cu at six concentrations, ranging from 6.9×10(-9) mol L(-1) to 1.0×10(-7) mol L(-1) free Cu(2+) ions. To guarantee the cells would be in a physiological status that reflected the external P concentration, they were previously acclimated up to constant growth rate at each P concentration. Phosphorus acclimated cells were then exposed to Cu and toxicity was evaluated through population density, growth rates and chlorophyll a content. Free Cu(2+) ions concentrations were calculated through the chemical equilibrium model MINEQL(+). The results showed that higher Cu toxicity was obtained in P-limited than in P-replete cells, and that chlorophyll a/cell was higher in P-limited cells and excess Cu than in P-replete cells. This confirms the importance of microalgae nutritional status to withstand the negative effects of the trace metal.