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Fermented swine urine (3%) (v/v) was added to a control medium (CT), named KEP I, and an aquatic microalgal culture (10% Bold's Basal Medium) for growing mixed Scenedesmus species. During a two-month period, the KEP I medium effected, the delayed onset of the stationary phase of cell division in a batch culture. After 31 days, of culturing, the growth rate (3-fold), dry weight (2.6-fold) and amino acid levels (2.7-fold), and secondary metabolites including chlorophyll a (2.1-fold), astaxanthin (2.8-fold), lutein (2.7-fold) and alpha (greater than 30-fold) and beta-carotene (greater than 5-fold) increased a greater degree in Scenedesmus grown in KEP I medium than in CT medium. Total lipids were much less in cells grown in KEP I than those grown in CT. An increased quantum yield of photosystem II of the aquatic microalgae. The KEP I medium should improve the cost efficiency of industrial mass batch cultures for CO(2) sequestration, bioremediation, phytonutrients, agricultural fertilizers, and microalgal stock for the species preservation of aquaculture strains for use in young fish feed. It may also serve to attenuate negative environmental impact via the recycling of animal wastewater.

Cd accumulation in cultures of Scenedesmus vacuolatus, a freshwater alga, was investigated in water samples from freshwater systems (two hard- and one softwater system) and compared to accumulation from synthetic culture medium. The Cd accumulation in S. vacuolatus was compared to total, labile, and free Cd measured by various analytical methods. In the hardwater samples, Cd accumulation was similar to accumulation from the culture medium and approximately followed the free ion activity model. In the softwater system, Cd accumulation increased with increasing free and total Cd concentrations, but was lower than expected from the free Cd2+ concentrations compared to the other freshwaters and culture media. Cd accumulation in the various freshwaters was not simply related to labile and free Cd concentrations measured by several methods. From experiments using fulvic acid as a ligand in the culture medium, no influence on Cd accumulation other than by modifying the Cd speciation was detected. Modeling of the Cd uptake revealed that besides cation and proton competition, other factors such as dissolved organic carbon composition and the presence of colloids might reduce Cd accumulation in softwater systems.

In order to disclosure the formation rule of predominant species in different nutrition conditions, three kinds of nutrition concentration were selected for the competition experiments with the common species of blue-green algae bloom Microcystis aeruginosa and the common species of green algae bloom Scenedesmus quadricauda. The competition relation was analysed by the competition parameters. The results indicate, in low nutrition, Scenedesmus quadricauda can stimulate the growth of Microcystis aeruginosa in mixed culture, the simulation becomes evident in low N/P ratio and M. aeruginosa can also stimulate the growth of S. quadricauda; in eutrophic condition, inhibition effect is connected with N/P; in hyper-eutrophic condition, the inhibition effect of S. quadricauda on M. aeruginosa is about three times as that of M. aeruginosa on S. quadricauda, and the effect of N/P ratio on competition inhibition parameters isn't evident. In low concentration N and P water, M. aeruginosa is easy to become predominant species, while in high concentration N and P water, S. quadricanda is easy to become predominant species.

The biodegradation of phenolic compounds by microalgae seems to be not a simple feature of a particular organism, but mostly a bioenergetic process depending on the growth conditions, especially on the exogenously supplied energy (carbon and light) sources. By using chlorophyll fluorescence induction measurements to estimate the molecular structure and function of the photosynthetic apparatus and therefore the tolerance/sensitivity of microalgae incubated with phenols, it can be assumed that, at least in low concentrations, phenol have no toxic effects on the cultures and can be used as alternative carbon source in them. Halophenols (chlorophenols, bromophenols and iodophenols) are quite toxic for the microalgal cultures. In halophenols the first step of the biodegradation is the split of the halogen substituent (dehalogenation). This is strongly determined by the bond dissociation energy of the corresponding substituent and therefore the energetic requirement for the biodegradation of halophenols increases following the sequence: iodophenol

The increase in the concentration of atmospheric carbon dioxide is considered to be one of the main causes of global warming. As estimated by the Intergovernmental Panel on Climate Change (IPCC) criteria, about 10-15% of the gases emitted from the combustion coal being in the form of carbon dioxide. Microalgae and cyanobacteria can contribute to the reduction of atmospheric carbon dioxide by using this gas as carbon source. We cultivated the Scenedesmus obliquus and Spirulina sp. at 30 degrees C in a temperature-controlled three-stage serial tubular photobioreactor and determined the resistance of these organisms to limitation and excess of carbon dioxide and the capacity of the system to fix this greenhouse gas. After 5 days of cultivation under conditions of carbon limitation both organisms showed cell death. Spirulina sp. presenting better results for all parameters than S. obliquus. For Spirulina sp. the maximum specific growth rate and maximum productivity was 0.44 d(-1), 0.22 g L(-1)d(-1), both with 6% (v/v) carbon dioxide and maximum cellular concentration was 3.50 g L(-1) with 12% (v/v) carbon dioxide. Maximum daily carbon dioxide biofixation was 53.29% for 6% (v/v) carbon dioxide and 45.61% for 12% carbon dioxide to Spirulina sp. corresponding values for S. obliquus being 28.08% for 6% (v/v) carbon dioxide and 13.56% for 12% (v/v) carbon dioxide. The highest mean carbon dioxide fixation rates value was 37.9% to Spirulina sp. in the 6% carbon dioxide runs.

In the present paper we investigated the effects of sub-lethal concentrations of Cu2+ in the growth and metabolism of Scenedesmus incrassatulus. We found that the effect of Cu2+ on growth, photosynthetic pigments (chlorophylls and carotenoids) and metabolism do not follow the same pattern. Photosynthesis was more sensitive than respiration. The analysis of chlorophyll a fluorescence transient shows that the effect of sub-lethal Cu2+ concentration in vivo, causes a reduction of the active PSII reaction centers and the primary charge separation, decreasing the quantum yield of PSII, the electron transport rate and the photosynthetic O2 evolution. The order of sensitivity found was: Growth>photosynthetic pigments content=photosynthetic O2 evolution>photosynthetic electron transport>respiration. The uncoupled relationship between growth and metabolism is discussed.

In the present study, Zn and Mn competition with Cd uptake was investigated in the freshwater alga Scenedesmus vacuolatus. Scenedesmus vacuolatus was exposed to experimental media with Cd and either Zn or Mn in short-term experiments; long-term experiments were undertaken to investigate the effect of growth on Cd accumulation. Cadmium accumulation in S. vacuolatus could be detected at very low exposure concentrations (free Cd2+, 2 x 10(-14) to 1 x 10(-11) M), and uptake was proportional to the free-Cd2+ concentration. Zinc was an effective competitive inhibitor of Cd uptake when the Zn2+ to Cd2+ ratio was greater than 14 in the exposure medium, whereas Mn competed with Cd for uptake above a Mn2+ to Cd2+ ratio of greater than 10,000. Binding constants for Cd and Zn affinity to the transport sites were determined (KZn and KCd). Values for KZn were slightly higher (log K = 9.4-9.8) than values for KCd (log K = 8.9-9.8). In contrast, Cd seemed not to compete with the Mn-binding sites for uptake over the Mn concentration range from 1 x 10(-10) to 1 x 10(-8) M. Determined values for the binding constants of Zn and Cd show that a simple model can be applied to predict Cd uptake at known Zn and Cd concentrations. The environmental implications of these results are discussed with respect to potential Cd toxicity for aquatic organisms.

Short-term (24h) experiments were performed to examine the effect of anthraquinone (ANTQ) and phenanthrenequinone (PHEQ) on two Scenedesmus armatus strains (B1-76 and 276-4d) grown in a batch culture system aerated with CO2 at a low (0.1%) or elevated (2%) concentration. ANTQ at concentrations within the range of 0.156-1.250 mg dm-3 inhibited the growth of B1-76 population in a concentration-dependent manner, and calculated EC50 for low-CO2 cells was 0.56 mg dm-3. The toxic effect of ANTQ on this strain was more pronounced in high-CO2 cells, where not only growth but also photosynthesis, respiration and SOD activity were significantly inhibited. In contrast, except for SOD activity, no ANTQ effects on strain 276-4d were found. PHEQ at concentrations within the range of 0.063-0.125 mg dm-3 inhibited the growth of B1-76 population in a concentration-dependent manner. The value of EC50 for low-CO2 B1-76 cells was 0.10 mg dm-3. PHEQ inhibited the growth of both strains regardless of CO2 concentration. In B1-76 cells affected by PHEQ, inhibition of photosynthesis was independent of the CO2 level, whereas the SOD activity was much higher in cultures aerated with 2% than with 0.1% CO2. Higher toxicity of PHEQ to strain 276-4d grown at 2% CO2 was accompanied by strong inhibition of photosynthesis, while in low-CO2 cells this process was slightly stimulated. The SOD activity in both low- and high-CO2 cells of strain 276-4d treated with PHEQ was 2-3 times higher compared with the controls. The pattern of SOD isoforms (PAGE analysis) obtained from cells exposed to ANTQ or PHEQ did not change compared with the controls, but the location of the SOD isoforms bands on gel was affected by the concentration of CO2. The results suggest that the strain-specific toxicity of ANTQ and PHEQ may result from oxidative stress. In addition, carbon dioxide appears to play an important role in the toxicity of quinones to algae.

This study was conducted to determine the role of algal-algal interactions in a multispecies microalgal population on their sensitivities to copper based on an enzyme inhibition assay using flow cytometric measures. Autofluorescence (chlorophyll a and phycocyanin) was used to identify species and count algal signals. The effect of multispecies population on copper toxicity of Microcystis aeruginousa was detected (1) at the same initial cell density, (2) at the same surface area, and (3) in the presence and absence of Chlorella pyrenoidosa and Scenedesmus obliquus. As copper concentrations increased, esterase activity of M. aeruginosa changed in a concentration-dependent manner. The 24 h EC(50) value of M. aeruginosa in the multispecies population was significantly (P < 0.05) higher than those in the single-species population. Compared with S. obliquus, the effect of C. pyrenoidosa on M. aeruginosa was more marked (the 24 h EC(50) value of copper on fluorescin diacetate fluorescence of M. aeruginosa was 11 microg/L). At 48 h copper exposure (6 microg/L) analysis of intracellular reactive oxygen species levels also showed similar algal-algal interactions in multispecies microalgal populations. The pigment assay suggested that these algal-algal interactions occurred only at low concentrations (< 13 microg/L, 24 and 48 h copper exposure). This study demonstrates the importance of using multispecies populations to estimate metal toxicity in natural waters.

Objective: To investigate the impact of various levels of sublethal temperature (26 degrees C, 31 degrees C, 33 degrees C, 36 degrees C, and 39 degrees C) on growth and heat shock protein (hsp) expression in freshwater green alga Scenedesmus quadricauda., Methods: Impact of selected levels of temperature on growth rate (based on optical density), population count, chlorophyll-a and biomass of the alga was evaluated in artificial growth medium for 19 days. To determine the induction of hsp in the alga, it was exposed to selected temperature levels for 3 h and further kept for 6 h at culturing condition at 26 degrees C. Induction of hsp was confirmed by immuno-detection followed by SDS-polyacrylamide gel electrophoresis., Results: The selected growth parameters such as growth rate, population count, chlorophyll-a and biomass were reduced significantly (P < 0.001) at 39 degrees C. However, hsp 70 expression was observed only at 39 degrees C., Conclusion: Temperature up to 36 degrees C may be considered as the limit of safe exposure for thermal stress for the alga Scenedesmus quadricauda.

This paper aims at the characterization of algogenic organic matter (AOM) produced by the cyanobacteria Anabaena flos-aqua and Microcystis aeruginosa and the green algae Scenedesmus quadricauda. Further, it is focused on the description of differences in the composition of extracellular organic matter (EOM) and intracellular organic matter (IOM), and on the demonstration of AOM affinity to aluminum and iron coagulants. The results from the conducted analyses imply a significant difference in the amount and properties of the proteins contained in EOM in comparison to IOM. The differences in the production of proteins also depend on the species of microorganism observed and its growth phase; ageing of the culture is accompanied by a gradual increase of the portion of proteins forming AOM. Using affinity chromatography (AC), the proteins with relative molecular weight around 60 kDa were isolated as a component of AOM of cyanobacteria A. flos-aqua and M. aeruginosa. These proteins are able to form complex compounds with iron and aluminum.

The color of light (white, red, blue, and green) had a significant effect on the growth and reproductive processes (both in the nucleocytoplasmic and chloroplast compartment of the cells) in synchronous cultures of Scenedesmus obliquus. This effect decreased in the order red > white > blue > green. In the same order, the light phase of the cell cycle (time when first autospores started to be released) was prolonged. The length of dark phase (time when 100 % of daughters were allowed to release from mothers) was not influenced and was the same for all colors. Critical cell size for cell division in green light was shifted to a smaller size (compared with cells grown in other lights) and so was the size of released daughters. The nuclear cycle was slowed in blue and even in green light, contrary to cells grown in red and white light. At the beginning of the cell cycle, one-nucleus daughters possess approximately 10 nucleoids; during the cell cycle their number doubled in all variants before the division of nuclei. Both events were delayed in cultures grown more slowly most markedly in green light. Smaller daughters in the green variant possessed a lower number of nucleoids. Motile cells released in continuous green or blue lights but not in red one were rarely observed.

Algae are exposed to elevated levels of heavy metals in water bodies generally for a long-term, and occasionally for a short-term duration. The present study deals with oxidative stress in Scenedesmus sp., commonly found in nutrient-rich freshwaters, during short- (6h) and long-term (7d) exposure to Cu(2+) and Zn(2+). The cells accumulated almost 2- and 4-times more Cu(2+) and Zn(2+) inside the cells during long-term than during short-term exposure to these metals. But the data on photosynthetic O(2) evolution and cell viability suggest that Scenedesmus sp. experienced lesser metal stress in long-term than in short-term experiment. Although malondialdehyde content was slightly higher in the long-term experiment, the amount produced by one unit intracellular metal was significantly lower than that in the short-term experiment. Superoxide dismutase activity of Scenedesmus sp. was >30% higher during long-term than during short-term exposure to Cu(2+) and Zn(2+). But, catalase and ascorbate peroxidase activities increased only at 2.5 microM Cu(2+) and 25 microM Zn(2+) when oxidative stress was mild, but were inhibited at 10 microM Cu(2+) under intense oxidative stress. Cu(2+) and Zn(2+) reduced glutathione reductase activity and total SH content of Scenedesmus sp. in both the experiments, with greater reduction occurring in the long-term experiment. The depletion of total thiol was positively related with the intracellular level of metals. Thiols might have helped Scenedesmus sp. in overcoming metal-induced oxidative stress, but depletion of thiol pool is known to make cells vulnerable to oxidative stress. The study suggests that antioxidant enzymes play a role only under mild oxidative stress. An increased accumulation of proline seems to be an important strategy for alleviating metal-induced oxidative stress in Scenedesmus sp. The study shows that Scenedesmus sp. could acclimatize during long-term exposure to toxic concentrations of the test metals.

Dissolved microcystins (MC) are regularly present in water dominated by microcystin-producing, bloom-forming cyanobacteria. In vitro experiments with environmentally feasible concentrations (5 x 10(-7) M) of the three most common microcystins, MC-LR, -RR, and -YR, revealed that they influence the metabolism of different representative phytoplanktons. At light intensities close to the cyanobacterial bloom environment (50 mumol m(-2) s(-1)), they produce morphological and physiological changes in both microcystin-producing and nonproducing Microcystis aeruginosa strains, and also have similar effects on the green alga Scenedesmus quadricauda that is frequently present in cyanobacterial blooms. All three microcystin variants tested induce cell aggregation, increase in cell volume, and overproduction of photosynthetic pigments. All three effects appear to be related to each other, but are not necessarily caused by the same mechanism. The biological activity of microcystins toward the light-harvesting complex of photobionts can be interpreted as a signal announcing the worsening of light conditions due to the massive proliferation of cyanobacteria. Although the function of microcystins is still unknown, it is evident that they have numerous effects on phytoplankton organisms in nature. These effects depend on the individual organism as well as on the various intracellular and extracellular signaling pathways. The fact that dissolved microcystins also influence the physiology of microcystin-producing cyanobacteria leads us to the conclusion that the role of microcystins in the producing cells differs from their role in the water environment.

Partial cDNAs corresponding to the GapA, GapC and GapN genes that encode the three different glyceraldehyde-3-phosphate dehydrogenases (GAPDHs) of the green microalga Scenedesmus vacuolatus SAG 211-8b have been cloned and characterized. Northern blot experiments, as well as immunoblots and activity measurements, demonstrate a differential regulation by sugars of the components of the algal Gap gene system. Addition of glucose or other metabolizable sugars to photoautotrophic cultures promoted a drastic repression of the GapA gene and depletion to negligible levels of the corresponding GAPDHA, a chloroplastic protein involved in photosynthetic CO2 assimilation. By contrast, expression of the GapC and GapN genes encoding their cytosolic counterparts involved in glycolysis was enhanced. However, no down-regulation of the GapA gene by glucose took place in the dark, indicating that the observed effect is associated with sugar assimilation in the light. Likewise, glucose promoted in illuminated algal cultures a severe decrease of photosystem II functionality, estimated by O2 evolution activity, thermoluminescence emission and D1 protein level, while again, no effect was observed in the dark. On the basis of the correlation found between photosystem II performance and sugar transcriptional regulation of the GapA gene, a scenario of sugar-mediated regulation of photosynthetic metabolism in microalgae is proposed that will help to explain the so-called glucose bleaching effect in photosynthetic eukaryotes.

The dynamics of the transition of V. cholerae into the uncultivable state in distilled, river and tap water, containing organic and inorganic components added, was studied. As additives, potassium nitrate, potassium phosphate, magnesium sulfate, ammonium chloride, lysine, alpha-ketoglutarate, succinic acid, catalase were used. The study of the influence of biotic factors on transition into the uncultivable state was carried out in the presence of one-celled green algae Scenedesmus quadricauda or infusoria Paramecium caudatum. The linear dependence of speed of transition into the uncultivable form on the concentration of cells was noted. The composition of the microcosmic medium was also found to have some influence on the speed of transition into the uncultivable form and on the reversibility of this process. The presence of organic substances, such as peptone solution or destroyed cells of phyto- and zooplankton, in the microcosmic medium prolonged the time of transition into the uncultivable form and produced a positive effect on the capacity of the population to reversion. In respect of live biotic components, no such dependence was found. Inorganic additives prolonged the time of transition into the uncultivable state, but did not promote reversion.

We examined the influences of three trace metals on the accumulation of a major nutrient (NO3-) in Scenedesmus quadricauda. A comparative study on metal-nutrient interaction in free and immobilized states of algal cells was conducted. The effect due to interaction between different variables (cell state type, metal type, and metal dose) was studied to assess the variation in the nitrate uptake by free and immobilized cells. The results analyzed by ANOVA (three-way) (components: cell state type, metal type, and metal dose) confirmed that the inhibition of nitrate uptake by test metals was highly significant (P<0.001). Free and immobilized states of S. quadricauda responded differently (P<0.05, ANOVA) to the types of metal added. Uptake kinetics was studied by monitoring short-term uptake rates at different nutrient levels. Free and immobilized cells of the organism displayed noncompetitive modes of inhibition for Ni and Zn while a competitive mode of inhibition by Cd was observed in both free and immobilized states of the organism.

Semicontinuous cultures of Scenedesmus obliquus in artificial wastewater were maintained with 30% and 40% daily dilutions and under a 14:10 h light-dark cycle, with temperatures of 25.5 and 17 degrees C during light and dark hours. Under this regime, the production of organic biomass was 39.3 and 25.2 mg l(-1)d(-1) for the 30% and 40% dilutions, and 24.9 and 16.7 mg l(-1)d(-1) of single-cell proteins. Most of the nitrogen removal took place during the light hours, with daily totals of 9.27 and 8.45 mg l(-1) for the 30% and 40% dilutions. With the former, 43.7% of the nitrogen removed was recycled by the microalgae into proteins and other organic nitrogen cell contents, but this efficiency decreased to 26.4% when the dilutions were raised to 40%.

This study examines the influences of Cd, Se, and Zn concentrations in ambient water on the assimilation, elimination, and release budget of metals, and the clearance rate of Daphnia magna through ingestion of phytoplankton diets. With increasing ambient metal concentration over two to three orders of magnitude, the assimilation efficiencies (AEs) of Cd and Se from two green algal species, Scenedesmus obliquus and Chlamydomonas reinhardtii, decreased significantly. In contrast, the Zn AE from ingested food, which typically was low (<7%), was not impacted by the difference in Zn or Cd concentrations in ingested cells. The Cd AE also was reduced significantly by a high Zn concentration in ingested algae, perhaps due to the competitive assimilation between the two metals. Metal elimination was determined after the daphnids were exposed to the radiolabeled diets for 3 d. The elimination rate constant was not dependent on the metal concentration in ingested food. Similarly, the routes of metal elimination (excretion, egestion, molting, and neonate production) were less influenced by the metal concentration. The rapid elimination of Se and Zn may be attributed to the transfer of these metals from mother to offspring. Except at the highest Cd (917 nM) and Se (1,333 nM) concentrations, there was no significant effect of metal concentration on the clearance rate of the animals. Consequently, the metal assimilation in D. magna may play an important role in the regulation of Cd and Se influx into the animals, whereas the regulation of the metal tissue burden in the animals is unlikely to be achieved through the metal elimination. The trophic transfer factors (TTF) of Cd and Se were related inversely to the metal concentration in the water, whereas the TTF of Zn was less variable at different Zn concentrations. The change of TTF with metal concentration will have important implication for ecotoxicity testing using D. magna, especially when the dietary exposure is incorporated.

An experimental investigation was carried out, and a mathematical model of interaction between invertebrates (infusoria Paramecium caudatum and rotifera Brachionus plicatilis) and algae (Chlorella vulgaris and Scenedesmus quadricauda) in the "producer-consumer" aquatic biotic cycle with spatially divided links was constructed. The model describes the dynamics of a mixed culture of infusoria and rotifera in the "consumer" link, when they consume a mixed culture of algae coming from the "producer" link. A negative influence of products of algae Scenedesmus metabolism upon the reproduction of infusoria P. caudatum was revealed. Taking this into account, a qualitative coincidence of the results of mathematical modeling with experimental data was obtained. It was shown that the co-existence of mixed algae culture in the "producer" link with invertebrates in the "consumer" link in the "producer-consumer" aquatic biotic cycle is impossible because of the displacement of infusoria P. caudatum by rotifera Brachionus plicatilis.

The sensitivity of different physiological parameters in Scenedesmus obliquus and Lemna minor exposed to herbicide (flumioxazin) was investigated to indicate the most convenient and sensitive parameter. To assess toxicity of flumioxazin, we used a panel of biomarkers: pigment contents, chlorophyll fluorescence parameters and antioxidative enzyme activities. Algae and duckweed were exposed to 48-h IC50 for growth rate. In L. minor, the sensitivity of the parameters was as follows: QN > Oxygen emmision > phiS(PSII) > QP > phi(PSII) > CAT, GR > Pigment> APO > Growth. For S. obliquus, this ranking was as follows: CAT > Oxygen emission > QP > APO > GR > Pigment > phiS(PSII) > Growth > phi(PSII) > QN (from the greatest to the least sensitive). The results demonstrated that the observed toxicity is related not only to interspecific variations but also to the selected parameter.

The microalga Scenedesmus incrassatulus was grown under continuous regime in the presence of chromium(VI), cadmium(II) and copper(II), as single metal species and as mixtures of two or three metals, in a laboratory scale system. We used an artificial wastewater with low free ion activities (as determined by MINEQL+) due to the presence of EDTA (a strong chelating agent) but with total concentrations not suitable for acceptable environments. Chromium(VI) and cadmium(II) had positive interaction that increased the removal percentages of both these metals; we could not, however, detect any interaction with copper(II). S. incrassatulus was able to remove all the tested metals to some extent (25-78%), but bivalent metals were not removed as efficiently as reported in batch cultures, probably due to the high pH values there recorded. Chromium(VI) was more efficiently removed in continuous cultures than in batch culture, because the uptake of chromate could be favored by actively growing algae., (Copyright 2004 Elsevier Ltd.)

Two strains of the unicellular green alga Scenedesmus acutus with different sensitivity to hexavalent chromium were compared for their tolerance of cadmium, by means of growth and recovery tests, and determination of cysteine, reduced glutathione and phytochelatin content, after short-term exposure to various cadmium concentrations (from 1.125 to 27 microM). Growth experiments showed that, after 7-day treatments with cadmium, the chromium-tolerant strain reached a significantly higher cell density and, after 24-h exposure to Cd, was able to resume growth significantly better than the wild type. Constitutive level of cysteine was higher in the chromium-tolerant strain, while glutathione levels were similar in the two strains. The higher content of cysteine and the maintenance of both reduced glutathione and phytochelatin high levels in the presence of cadmium, support the higher cadmium co-tolerance of the chromium-tolerant strain in comparison with the wild type one., (Copyright 2003 Elsevier B.V.)

A 4-h exposure of Scenedesmus sp. to Cu or Zn enhanced intracellular levels of both test metals and proline. The level of intracellular proline increased markedly up to 10 microM Cu, but higher concentrations were inhibitory. However, intracellular proline consistently increased with increasing concentration of Zn in the medium. Cu and Zn induced oxidative stress in the test alga by increasing lipid peroxidation and membrane permeability, and by reducing SH content. Pretreatment of the test alga with 1 mM proline for 30 min completely alleviated Cu-induced lipid peroxidation, minimized K+ efflux and also reduced depletion of the SH pool. But proline pretreatment could only slightly reduce Zn-induced oxidative stress. Interestingly, proline pretreatment increased the level of Cu (25-54%) and Zn (19-49%) inside the cells. It did not affect the activities of superoxide dismutase, ascorbate peroxidase or catalase, but improved glutathione reductase activity under Cu and Zn stress. A comparison of the effects of proline pretreatment on lipid peroxidation by Cu, Zn, methyl viologen and ultraviolet-B radiation suggests that proline protects cells from metal-induced oxidative stress by scavenging reactive oxygen species rather than by chelating metal ions. Pretreatment of cells with a known antioxidant (ascorbate) and a hydroxyl radical scavenger (sodium benzoate) considerably reduced metal-induced lipid peroxidation and proline accumulation. However, sodium benzoate had a very mild effect on Zn-induced lipid peroxidation and proline accumulation. The present study demonstrates that proline possibly acts by detoxifying reactive oxygen species, mainly hydroxyl radicals, rather than by improving the antioxidant defense system under metal stress.

Many abiotic and biotic factors can influence the partitioning equilibrium of heavy metals, thus influencing metal impact on aquatic environments. Unicellular algal species release soluble organic substances able to complex metals. In our laboratory a Cr-tolerant strain was selected and isolated from a wild type strain of Scenedesmus acutus. The exudates released by the two strains counteracted the growth inhibition caused by Cr(VI) and the exudates of the Cr-tolerant strain were more effective. On the contrary, the exudates did not reduce chromium toxicity to the cladoceran Daphnia magna. The reduction of chromium effect on algae seems the consequence of an algae-specific interaction among Cr(VI), exudates and algal cells. Chromium uptake resulted to be energy-dependent since bioaccumulation rate in subdued light condition was lower than at high light intensity. The effect of Cr(VI) on algae changed depending on metabolism of the cells and in particular it seemed to be related to the bioaccumulation rate. Tolerance in the selected strain could not be ascribed to a lower uptake of chromium. The difference in sensitivity to chromium between the two strains was exploited to evaluate if tolerance acquired by algae could have consequences for Daphnia. After treatment with Cr(VI), the two strains of S. acutus were used as food source for D. magna. The results indicate that chromium is accumulated by algae in a form not available for daphnids and that Cr tolerance acquired by the algae can be of some advantage to the consumer organism.

The uptake of Cd, Se(IV) and Zn by the freshwater alga Scenedesmus obliquus and the subsequent transfer and release budget in Daphnia magna were investigated under different nutrient additions and cell incubation conditions. An increase in ambient phosphate concentrations from 0.5 micromol l(-1) to 50 micromol l(-1) significantly increased the intracellular accumulation of Cd (by 18x) and Zn (by 5x), but decreased the accumulation of Se (by 126x) in the alga. The percentage of these metals distributing in the intracellular pool of algae also increased substantially with increasing ambient P concentrations. Nitrate addition from 5.0 to 200 micromol l(-1) did not influence the uptake of any of the three metals, although a significant decrease in the intracellular Se distribution was observed. Radiolabeled algae under different nutrient manipulations (semi-continuous culture, starvation, and P-pulse treatments) were used to measure trophic transfer assimilation efficiency (AE) in Daphnia. When the algal cells were grown in a semi-continuous culture, starved for N and P, or were treated with P-pulse, the AEs of Cd and Zn were generally independent of the nutritional conditions, but the Se AE was significantly affected by different P levels. The efflux rate constants, determined during 10 d depuration following 7 days of dietary uptake, decreased significantly for Cd and Zn, but were relatively constant for Se with increasing P concentration. N-addition caused no effect on the metal efflux rate constants. P- or N-additions did not influence the release budget (including molting, neonates, excretion and feces) for all three elements in Daphnia. Our study indicated that phosphate enrichment may substantially increase metal uptake in green alga S. obliquus. Responses of trophic transfer in Daphnia to nutrient enrichment were metal specific. P-enrichment can possibly lead to considerable decrease on Se transfer from algae to zooplankton.

A study of the effects of elevated levels of Cu2+ and Zn2+ on NO3- uptake and nitrate reductase (NR) activity in Scenedesmus sp. was carried out. The two metals inhibited NR and NO3- uptake in a concentration-dependent manner, with the latter process being inhibited more strongly than the former. After withdrawal of metal stress, NR activity and NO3- uptake recovered in a metal ion concentration-dependent manner. Dark pretreatment of the alga enhanced the toxic effects of the metal ions on NR activity and NO3- uptake. The recovery from metal stress was slower in the dark-pretreated cells in comparison to the light-pretreated cells. No recovery of NR and NO3- uptake occurred in the presence of the photosynthetic inhibitor, 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU), suggesting that photosynthesis was required for the recovery from metal stress. Cycloheximide blocked the recovery of NR activity in metal-treated alga, suggesting that new enzyme synthesis was required for the recovery from metal stress.

In crustaceans, cholesterol is an essential nutrient, which they must directly obtain from their food or by bioconversion from other dietary sterols. Eukaryotic phytoplankton contain a great variety of sterols that differ from cholesterol in having additional substituents or different positions and/or number of double bonds in the side chain or in the sterol nucleus. In this study, we investigated to what extent these structural features affect the growth and reproduction of Daphnia galeata in standardized growth experiments with the cyanobacterium Synechococcus elongatus supplemented with single sterols as food source. The results indicated that delta 5 (sitosterol, stigmasterol, desmosterol) and delta 5.7 (7-dehydrocholesterol, ergosterol) sterols meet the nutritional requirements of the daphnids, while the delta 7 sterol lathosterol supports somatic growth and reproduction to a significantly lower extent than cholesterol. Dihydrocholesterol (delta 0) and lanosterol (delta 8) did not improve the growth of D. galeata, and growth was adversely affected by the delta 4 sterol allocholesterol. Sterols seem to differ in their allocation to somatic growth and reproduction. Thus, structural differences of dietary sterols have pronounced effects on life-history traits of D. galeata.

A thioredoxin-like chloroplast protein of the fructosebisphosphatase-stimulating f-type, but with an unusually high molecular mass of 28 kDa has previously been identified and purified to homogeneity in a fractionation scheme for resolution of the acid- and heat-stable, regular-size (12kDa) thioredoxins of the unicellular green algae, Scenedesmus obliquus. An apparently analogous protein of 26 kDa was described in a cyanobacterium, Anabaena sp., but no such large thioredoxin species f exists in the thioredoxin profiles of higher plants. The structure of the 28 kDa protein, which had been envisaged to represent a precursor, or fusion product of the two more specialized, common chloroplast thioredoxins f and m has now been determined by amino acid sequencing. Although it exhibits virtually all the properties and enzyme-modulating activities of a thioredoxin proper this algal protein, surprisingly, does not belong to the thioredoxin family of small redox proteins but is identical with OEE (oxygen evolving enhancer) protein 1, an auxiliary component of the photosystem II manganese cluster. Extracts of Chlorella vulgaris and Chlamydomonas reinhardtii also contain heat-stable protein fractions of 23-26 kDa capable of specifically stimulating chloroplast fructosebisphosphatase in vitro. In contrast, OEE protein 1 from spinach is not able to modulate FbPase or NADP malate dehydrogenase from spinach chloroplasts. A dual function of the OEE protein in algal photosynthesis is envisaged.

Plastocyanin and cytochrome c6 from the green alga Scenedesmus vacuolatus were immunoquantified in cells grown under different concentrations of copper and iron. Plastocyanin expression was constitutive, its synthesis was not significantly affected by iron availability, and increases with copper availability. On the contrary, cytochrome c6 synthesis is repressed by copper, and only residual amounts of the protein were detected at 0.1 micromol/L copper. Under copper deficiency, cytochrome c6 is slightly dependent on iron. In natural environments, plastocyanin seems to be the predominant electron donor to P700.

Little is known about the fate and the effects of phthalic acid monoesters. Various of these monoesters ranging from n-butyl to isononyl monoester have been evaluated in respect to their biodegradation behaviour and their acute aquatic toxicity. All esters are readily biodegradable, achieving degradation rates of 90% and more. The acute toxicity values strongly depend on the carbon chain length of the alcohol moiety. The short chain specimen have LC/EC50 around and above 100 mg/l, with values levelling off to around 30 mg/l for the isononyl monoester.

The time course of microsecond-delayed light emission and that of prompt fluorescence was measured during the period of light adaptation for the wild-type strain and mutants Nos 8 and 11 of Scenedesmus obliquus, Strain D3, exposed t stimulus light in a photon-counting apparatus. As compared to prompt fluorescence, the delayed light emission was generally more susceptible to change in experimental conditions. (1) Illuminating normally grown wild-type cells with higher light intensities (100 mW/cm2 or higher), the intensity of delayed light increases to a maximum in th first few minutes, then decreases to a steady-state level after about 1 h. The maximu is missing at lower exciting light intensities. (2) Illuminating dark-adapted wild-type cells with high light intensities, the time course of delayed light emission is similar to that one described above, apart from a lag period within the very first minutes of light adaptation. That lag period is characterized by a relative minimum of the intensity lying higher than the intensity of the steady-state level. The length and the profile of the lag period depends on the time of dark adaptation. With increasing dark adaptation time the maximum of delayed light intensity appears later. (3) This maximum of delayed light intensity is only present for times shorter than 100 ps after turning off the stimulus light. (4) For mutant No. 8 the intensity of delayed light shows a steady decrease during the time course. (5) For light-adapted wild-type cells the intensity of delayed light and that of prompt fluorescence increases linearly with increasing intensity of exciting light below the saturation point in the light curve (about 10 mW/cm2). The increase is less than linear above the saturation point. The results are discussed in terms of two photosystems coupled by an electron transfer chain.