Bioaccumulation of metals by algae from acid mine drainage: a case study of Frongoch Mine (UK).

In Frongoch Mine (UK), it is unclear the distribution of metals on indigenous algae and whether these species of algae can accumulate metals. This study aimed to investigate the role of indigenous algae for metal removal from acid mine drainage and understand if metals can be adsorbed on the surface of algae or/and bioaccumulated in algae. A sequential extraction procedure was applied for algae samples collected from acid mine drainage (AMD) water to identify the forms in which metals are found in algae. Concentrations of Fe, Pb, Zn, Cu and Cd were evaluated in the algae and AMD samples were collected in June and October 2019. AMDs samples had a pH value ranging between 3.5 and 6.9 and high concentrations of Zn (351 mg/L) and Pb (4.22 mg/L) that exceeded the water quality standards (Water Framework Directive, 2015). Algae Ulothrix sp. and Oedogonium sp. were the two main species in the Frongoch AMDs. The concentrations of metals in algae ranged from 0.007 to 51 mg/g, and the bioconcentration factor of metals decreased in the following order: Fe > > Pb > > Cu > Cd > Zn. It was found that Zn, Cu and Cd were adsorbed onto the surface of and bioaccumulated in the algae, while Pb and Fe were mainly bioaccumulated in the algae. Indigenous algae can be considered as a biogeochemical barrier where metals are accumulating and can be used in bioremediation methods. Also, indigenous algae could be used as a bioindicator to assess water pollution at Frongoch Mine and other similar metal mines.

The effects of copper on photosynthesis and biomolecules yield in Chlorolobion braunii.

Our knowledge of the effects of copper on microalgal physiology is largely based on studies conducted with high copper concentrations; much less is known when environmentally relevant copper levels come into question. Here, we evaluated the physiology of Chlorolobion braunii exposed to free copper ion concentrations between 5.7 × 10-9 and 5.0 × 10-6  mol · L-1 , thus including environmentally relevant values. Population growth and maximum photosynthetic quantum yield of PSII were determined daily during the 96 h laboratory controlled experiment. Exponentially-growing cells (48 h) were analyzed for effective quantum yield and rapid light curves (RLC), and total lipids, proteins, carbohydrates, chlorophyll a and carotenoids were determined. The results showed that growth rates and population density decreased gradually as copper increased in experiment, but the photosynthetic parameters (maximum and effective quantum yields) and photochemical quenching (qP) decreased only at the highest free copper concentration tested (5.0 × 10-6 mol · L-1 ); nonphotochemical quenching (NPQ) increased gradually with copper increase. The RLC parameters Ek and rETRmax were inversely proportional to copper concentration, while α and Im decreased only at 5.0 × 10-6 mol · L-1 . The effects of copper in biomolecules yield (mg · L-1 ) varied depending on the biomolecule. Lipid yield increased at free copper concentration as low as 2.5 × 10-8 mol · L-1 , but proteins and carbohydrates were constant throughout.

Cu and Cd affect distinctly the physiology of a cosmopolitan tropical freshwater phytoplankton.

Copper and Cd are natural constituents of freshwater ecosystems, both cycling influenced by microbial communities. The present research examined the impacts of environmentally relevant concentrations of Cu and Cd on the growth, viability, cell size, chlorophyll a (Chl a) content and photochemical efficiency of the tropical freshwater phytoplankton Chlorolobion braunii. Cell growth was significantly impaired by Cu and Cd, with EC50 occurring at 33.6 and 1.6µM, respectively. At sublethal levels (< EC50), cell death was already induced at 5µM Cu and 1µMCd. Average cell volume significantly increased as metal concentrations increased, as did the Chl a content per cell, although the Chl a content per unit volume decreased. Copper did not affect both the photosystem II (PSII) maximum quantum yield (ΦM) or the operational quantum yield (ΦE), while Cd significantly impacted ΦE, with EC50 occurring at 18.4µM. Different responses for Cu and Cd were obtained whether the photochemical fluorescence quenching (Qp) or non-photochemical quenching (Qn) were considered. Qp decreased after Cd addition, but was not altered after Cu addition. Qn values significantly increased after the addition of either metal. Non-photochemical quenching due to heat dissipation (NPQ) significantly increased in response to both metals, but it was more pronounced in the case of Cd. Overall, Cd was more toxic to C. braunii than Cu.

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.

Microalgae population dynamics in photobioreactors with secondary sewage effluent as culture medium.

Nitrogen and phosphorus present in sewage can be used for microalgae growth, possibiliting cost reduction in the production of microalgae at the same time that it decreases the eutrophication potential of the effluent. This research aimed at monitoring the native community of microalgae and coliform bacteria in a secondary effluent from anaerobic municipal sewage treatment. Two treatments (aerated and non-aerated) were performed to grow microalgae under semi-controlled conditions in semi-closed photobioreactors in a greenhouse. The results showed no significant pH and coliforms (total and Escherichia coli ) variation between treatments. Nutrient concentrations were reduced supporting microalgae growth up to 10 (7) cells.mL (-1) independent of aeration. Exponential growth was obtained from the first day for the non-aerated, but a 5 day lag phase of growth was obtained for the aerated. Chlorella vulgaris was the dominant microalgae (99.9%) in both treatments. In the aerated, 5 algae classes were detected (Chlorophyceae, Cyanophyceae, Chrysophyceae, Bacillariophyceae and Euglenophyceae), with 12 taxa, whereas in the non-aerated, 2 classes were identified (Chlorophyceae and Cyanophyceae), with 5 taxa. We concluded that effluent is viable for microalgae growth, especially Chlorella vulgaris, at the same time that the eutrophication potential and coliforms are decreased, contributing for better quality of the final effluent.

Microalgae population dynamics in photobioreactors with secondary sewage effluent as culture medium

Nitrogen and phosphorus present in sewage can be used for microalgae growth, possibiliting cost reduction in the production of microalgae at the same time that it decreases the eutrophication potential of the effluent. This research aimed at monitoring the native community of microalgae and coliform bacteria in a secondary effluent from anaerobic municipal sewage treatment. Two treatments (aerated and non-aerated) were performed to grow microalgae under semi-controlled conditions in semi-closed photobioreactors in a greenhouse. The results showed no significant pH and coliforms (total and Escherichia coli) variation between treatments. Nutrient concentrations were reduced supporting microalgae growth up to 107 cells.mL−1 independent of aeration. Exponential growth was obtained from the first day for the non-aerated, but a 5 day lag phase of growth was obtained for the aerated. Chlorella vulgaris was the dominant microalgae (99.9%) in both treatments. In the aerated, 5 algae classes were detected (Chlorophyceae, Cyanophyceae, Chrysophyceae, Bacillariophyceae and Euglenophyceae), with 12 taxa, whereas in the non-aerated, 2 classes were identified (Chlorophyceae and Cyanophyceae), with 5 taxa. We concluded that effluent is viable for microalgae growth, especially Chlorella vulgaris, at the same time that the eutrophication potential and coliforms are decreased, contributing for better quality of the final effluent.

Influence of the microbial loop on trophodynamics and toxicity of cadmium complexed by cyanobacterium exudates.

Cadmium (Cd) is an important industrial and toxic metal. Its fate and toxicity in the environment may be mediated by association with dissolved organic materials excreted by phytoplankton. The aim of the present study was to investigate the trophodynamics and toxicity of Cd complexed with Cylindrospermopsis raciborskii exudates in a plankton food chain. The microbial loop involves heterotrophic bacteria as the primary consumer, which is supplemented with a Cd-exudate complex. The secondary consumer (protozoan Paramecium caudatum) was fed on the bacteria, and the tertiary consumer (copepod Mesocyclops longisetus) on the protozoa. The Cd complexing properties of the exudate were determined before the experiments, to ensure that all Cd was furnished to the organisms as the organic complex alone. The results showed that free Cd2+ ions caused the highest observed toxicity to bacteria, while Cd complexed to the cyanobacterial exudate was less toxic, but could be bioavailable and transferred through the food web. This study is a contribution to aquatic ecosystem management and to current knowledge of Cd dynamics, bioavailability and interaction with aquatic planktonic organisms.

Growth and biochemical composition of Chlorella vulgaris in different growth media.

The need for clean and low-cost algae production demands for investigations on algal physiological response under different growth conditions. In this research, we investigated the growth, biomass production and biochemical composition of Chlorella vulgaris using semi-continuous cultures employing three growth media (LC Oligo, Chu 10 and WC media). The highest cell density was obtained in LC Oligo, while the lowest in Chu medium. Chlorophyll a, carbohydrate and protein concentrations and yield were highest in Chu and LC Oligo media. Lipid class analysis showed that hydrocarbons (HC), sterol esthers (SE), free fatty acids (FFA), aliphatic alcohols (ALC), acetone mobile polar lipids (AMPL) and phospholipids (PL) concentrations and yields were highest in the Chu medium. Triglyceride (TAG) and sterol (ST) concentrations were highest in the LC Oligo medium. The results suggested that for cost effective cultivation, LC Oligo medium is the best choice among those studied, as it saved the cost of buying vitamins and EDTA associated with the other growth media, while at the same time resulted in the best growth performance and biomass production.

The effects of copper on the photosynthetic response of Phaeocystis cordata.

We investigated the effects of limiting (1.96 × 10(-9) mol l(-1) total Cu, corresponding to pCu 14.8; where pCu = -log [Cu(2+)]) and toxic Cu concentrations up to 8.0 × 10(-5) mol l(-1) total Cu (equivalent to pCu 9.5) on growth rates and photosynthetic activity of exponentially grown Phaeocystis cordata, using batch and semi-continuous cultures. With pulse amplitude modulated (PAM) fluorometry, we determined the photochemical response of P. cordata to the various Cu levels, and showed contrasting results for the batch and semi-continuous cultures. Although maximum photosystem II (PSII) quantum yield (Φ(M)) was optimal and constant in the semi-continuous P. cordata, the batch cultures showed a significant decrease in Φ(M) with culture age (0-72 h). The EC50 for the batch cultures was higher (2.0 × 10(-10) mol l(-1), pCu9.7), than that for the semi-continuous cultures (6.3 × 10(-11) mol l(-1), pCu10.2). The semi-continuous cultures exhibited a systematic and linear decrease in Φ(M) as Cu levels increased (for [Cu(2+)] < 1.0 × 10(-12) mol l(-1), pCu12.0), however, no effect of high Cu was observed on their operational PSII quantum yield (Φ'(M)). Similarly, semi-continuous cultures exhibited a significant decrease in Φ(M), but not in Φ'(M), because of low-Cu levels. Thus, Cu toxicity and Cu limitation damage the PSII reaction centers, but not the processes downstream of PSII. Quenching mechanisms (NPQ and Q (n)) were lower under high Cu relative to the controls, suggesting that toxic Cu impairs photo-protective mechanisms. PAM fluorometry is a sensitive tool for detecting minor physiological variations. However, culturing techniques (batch vs. semi-continuous) and sampling time might account for literature discrepancies on the effects of Cu on PSII. Semi-continuous culturing might be the most adequate technique to investigate Cu effects on PSII photochemistry.

Biological leaching of Mn, Al, Zn, Cu and Ti in an anaerobic sewage sludge effectuated by Thiobacillus ferrooxidans and its effect on metal partitioning.

The chemical fractionation and bioleaching of Mn, Al, Zn, Cu and Ti in municipal sewage sludge were investigated using Thiobacillus ferrooxidans as leaching microorganism. As a result of the bacterial activity, ORP increase and pH reduction were observed. Metal solubilization was accomplished only in experimental systems supplemented with energy source (Fe(II)). The solubilization efficiency approached approximately 80% for Mn and Zn, 24% for Cu, 10% for Al and 0.2% for Ti. The chemical fractionation of Mn, Al, Zn, Cu and Ti was investigated using a five-step sequential extraction procedure employing KNO3, KF, Na4P2O7, EDTA and HNO3. The results show that the bioleaching process affected the partitioning of Mn and Zn, increasing its percentage of elution in the KNO3 fraction while reducing it in the KF, Na4P2O7 and EDTA fractions. No significant effect was detected on the partitioning of Cu and Al. However, quantitatively the metals Mn, Zn, Cu and Al were extracted with higher efficiency after the bacterial activity. Titanium was unaffected by the bioleaching process in both qualitative and quantitative aspects.