The use of herbicides constitutes the principal method of weed control but the introduction of these compounds into the aquatic environment can provoke severe consequences for non-target organisms such as microalgae. Toxic effects of these pollutants on microalgae are generally evaluated using phytotoxicity tests based on growth inhibition, a population-based parameter. However, physiological cellular endpoints could allow early detection of cell stress and elucidate underlying toxicity mechanisms. Effects of the herbicide paraquat on the freshwater microalga Chlamydomonas moewusii were studied to evaluate growth rate and cellular parameters such as cellular viability and metabolic activity assayed by flow cytometry and DNA damage assayed by the comet assay. Sensitivity of growth and parameters assayed by flow cytometry were similar, showing a significant effect in cultures exposed to a paraquat concentration of 0.1 microM or higher, although in cultures exposed during 48 h to 0.05 microM, a significant stimulation of cellular fluorescein fluorescence was observed, related to cellular metabolic activity. After only 24 h of herbicide exposure significant DNA damage was observed in microalgal cells exposed to all paraquat concentrations assayed, with a 23.67% of comets in cultures exposed to 0.05 microM, revealing the genotoxicity of this herbicide. Taking into account the results obtained, comet assay provides a sensitive and rapid system for measuring primary DNA damage in Chlamydomonas moewusii, which could be an important aspect of environmental genotoxicity monitoring in surface waters.
The effects of two herbicides used wide-spread, isoproturon (phenylurea) and terbutryn (triazine), on growth, dry weight, elemental composition, photosynthetic pigments and protein content, and cell volume assayed by flow cytometry in the freshwater microalgae Chlorella vulgaris were studied. Different parameters for algal activity show widely different sensitivities to these aquatic pollutants. After 96 h of herbicide exposure, terbutryn was the strongest inhibitor of growth, giving an EC50 value for growth twice lower than that for isoproturon cultures (EC50 terbutryn=0.097 microM; EC50 isoproturon=0.199 microM). However, lower concentrations of the triazine herbicide provoked an increase in the cellular density and growth rate of this microalga, not observed in the phenylurea-treated cultures. Cellular volume and dry weight of C. vulgaris cells were increased strongly in the presence of isoproturon and terbutryn. Other cellular parameters, such as pigment and protein content, were stimulated with both herbicides at higher concentrations.
Investigation of herbicide toxicology in non-target aquatic primary producers such as microalgae is of great importance from an ecological point of view. In order to study the toxicity of the widely used herbicide paraquat on freshwater green microalga Chlamydomonas moewusii, physiological changes associated with 96 h-exposures to this pollutant were monitored using flow cytometry (FCM) technique. Intracellular reactive oxygen species concentration, cytoplasmic membrane potential, metabolic activity and cell protein content were monitored to evaluate the toxicological impact of paraquat on algal physiology. Results showed that herbicide paraquat induced oxidative stress in C. moewusii cells, as it indicated the increase of both superoxide anion and hydrogen peroxide levels observed in non-chlorotic cells of cultures exposed to increasing herbicide concentrations. Furthermore, a progressive increase in the percentage of depolarised cells and a decrease in the metabolic activity level were observed in response to paraquat when non-chlorotic cells were analysed. Chlorotic cells were probably non-viable cells, based on the cytoplasmic membrane depolarisation, its metabolically non-active state and its drastically reduced protein content. In view of the obtained results, we have concluded that a range of significant physiological alterations, detected by flow cytometry, occur when C. moewusii, an ubiquitous microalga in freshwater environments, is challenged with environmentally relevant concentrations of the herbicide paraquat.
Aquaculture facilities are a potential source of antibiotics to the aquatic ecosystems. The presence of these compounds in the environment may have deleterious effects on non-target aquatic organisms such as microalgae, which are often used as biological indicators of pollution. Therefore, the toxicity induced by chloramphenicol (CHL), florphenicol (FLO) and oxytetracycline (OTC), three antibiotics widely used in aquaculture, on the marine microalga Tetraselmis suecica was evaluated. Growth inhibition and physiological and biochemical parameters were analysed. All three antibiotics inhibited growth of T. suecica with 96 h IC50 values of 11.16, 9.03 and 17.25 mg L(-1) for CHL, FLO and OTC, respectively. After 24 h of exposure no effects on growth were observed and cell viability was also unaffected, whereas a decrease in esterase activity, related with cell vitality, was observed at the higher concentrations assayed. Photosynthesis related parameters such as chlorophyll a cellular content and autofluorescence were also altered after 24 h of antibiotics addition. It can be concluded that T. suecica was sensitive to the three antibiotics tested.
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