Assessment of water quality in Aguieira reservoir: Ecotoxicological tools in addition to the Water Framework Directive.

To assess the ecological potential (EP), the Water Framework Directive (WFD) uses specific parameters but ignores ecosystem functioning and individual/subindividual biological responses. The heavily modified water bodies (reservoirs) are strongly influenced by human activities that promote their imbalance, with a loss or compromise the biodiversity and the functioning of these ecosystems. This work aims to determinate the EP according physical, chemical and biological parameters of WFD and evaluate the functionality and sensitivity of ecotoxicological tools in the evaluation of water quality of Aguieira reservoir. Four sampling sites were defined in Aguieira reservoir and sampling was conducted in autumn 2018, spring and autumn 2019. WFD showed overall that Aguieira reservoir presented moderate EP. The autumn samples and the two reservoir upstream sites had the worst classification. These results were complemented by a set of ecotoxicological assays (growth inhibition of Raphidocelis subcapitata and Spirodela polyrhiza, and feeding rate of Daphnia longispina) using three natural water treatments. R. subcapitata was the most sensitive species, indicating the presence of a risk that is not detected by chemical analysis, nor by organisms of different trophic levels. D. longispina only showed sensitivity in A3 and A4 in autumn, however this results was difficult to interpret since no reason can be observed to respond the F1 lower feeding rate observed in almost sites. S. polyrhiza, as well as the total chlorophyll, lipid peroxidation, and proline content did not show sensitivity, since no discrimination among the water treatments were recorded. The relevance of bioassays is associated with the fact that an integrated response to a complex mixture can be obtained, which can contain more substances than the measured ones. Thus, we may conclude that WFD are insufficient to draw conclusions about water quality, and the growth inhibition of Raphidocelis subcapitata assay is an important parameter to complement the WFD. In addition, species of different trophic levels are recommended for ecotoxicological analyses due to differences in species sensitivities.

Adequacy of planctomycetes as supplementary food source for Daphnia magna.

The nutritional quality of daphnids diet can influence their growth, reproduction and survival. In aquatic ecosystems, bacteria can contribute significantly to Daphnia diet by supporting, for instances, their high needs for phosphorus. The laboratory feeding of the model organisms Daphnia spp. is algal based, but should be improved to allow their better performance. The aim of this study was to evaluate the potential of two planctomycetes, Gemmata obscuriglobus and Rhodopirellula rubra, from exponential and stationary growth phases as alternative or supplementary food source for Daphnia magna. The actinobacterium Arthrobacter sp. was used for comparison. The feeding with only bacteria showed the inefficacy of both planctomycetes and actinobacteria as the only food source. However, when used in supplement to Raphidocelis subcapitata, a decrease in the age of first reproduction, a significant increase in reproductive output, in somatic growth and in rate of population increase was found for the highest cell densities of bacteria tested. The typical pink coloration of these bacteria present in daphnids body and eggs confirmed bacterial absorption and metabolization of their pigment. Planctomycetes yielded better results than the actinobacteria Arthrobacter but G. obscuriglobus that possesses sterols did not induce a better performance comparatively to R. rubra. No relation could be established between the feeding treatments that allowed improvement of Daphnia performance and the different kind of Daphnia' Fatty Acid Methyl Esters. The use of sonication to separate planctomycetal cells before feeding the daphnids proved to be efficient. We confirmed that R. subcapitata supplemented by bacteria allows a better growth performance of D. magna.

Flow cytometric analysis of chronic and acute toxicity of copper(II) on the marine dinoflagellate Amphidinium carterae.

BACKGROUND: Copper(II) is a heavy metal whose levels have increased in some marine ecosystems to polluting levels. Dinoflagellates, an important phytoplankton group, are at the base of aquatic food chains and bioaccumulation of copper by these microorganisms can result in complex ecosystem alterations, so we investigated how copper disturbs those cells. METHODS: Cytotoxic effects of sublethal and lethal copper concentrations ranging from 4.2 nM (control condition) to 3.13 microM estimated labile copper were studied in batch cultures of Amphidinium carterae. Cell morphology, motility, autofluorescence, and fluorescein diacetate (FDA)-dependent fluorescence generation were evaluated by flow cytometry (FCM) and microscopy. RESULTS: Exposure of A. carterae to toxic levels of copper impaired cell mobility, delayed cell proliferation, led to increased green autofluorescence, and at 3.13 microM labile copper also induced encystment and death. Chlorophyll fluorescence, however, was not affected. Kinetic FCM assay of FDA-dependent fluorescence generation showed a dose-dependent enhancement of fluorescein fluorescence immediately after copper addition and in cultures with sustained exposure to this toxicant. CONCLUSIONS: Our data suggest that copper toxicity occurs quickly at the membrane level in relation to oxidative stress generation. Based on fluorescence kinetic studies, the Na(+)/H(+) antiporter seemed to be affected by copper, thereby affecting intracellular pH.

Electrochemical evidence of surfactant activity of the hepes pH buffer which may have implications on trace metal availability to cultures in vitro.

Hydrogen ion buffers are required for many different types of in vitro biological and chemical studies, but they may not be inert enough, thus interfering with the system under study. N-Hydroxyethylpiperazine-N'-ethanesulfonic acid (Hepes), a zwitterionic buffer in the 7.0-8.0 pH range, has been used in different biological studies. In this work, surfactant activity of Hepes and the buffer's capability to bind copper(II) were investigated through different electrochemical techniques: ion-selective electrode potentiometry, potentiometric stripping analysis, differential pulse anodic stripping voltammetry, normal pulse anodic stripping voltammetry, and alternating current polarography. Hepes concentrations between 0.1 (0.24 g liter-1) and 25 mmol liter-1 (6.0 g liter-1) were studied at pH 8.0 (and at pH 6.5 for comparison). At pH 8.0 Hepes displayed surfactant activity, particularly when magnitude of Hepes > 10 mmol liter-1 (2.4 g liter-1). The ability of Hepes to bind copper(II) was detected when Hepes was in large excess (magnitude of Hepes/magnitude of Cu between 10,000 and 2000). Eventual implications in the bioavailability of copper(II) are also discussed.