Sub-lethal combined effects of illicit drug and decreased pH on marine mussels: A short-time exposure to crack cocaine in CO2 enrichment scenarios.

The increasing CO2-concentrations in the atmosphere promote ocean acidification. Seawater chemistry changes interact with contaminants, such as illicit drugs in the coastal zones. This work evaluates impacts of pH decrease and crack-cocaine exposure on the commercial mussel Perna perna through biomarker responses (lysosomal membrane stability, lipid peroxidation, and DNA strand breaks). The organisms were exposed to different crack-cocaine concentrations (0.5, 5.0, and 50 µg L-1) combined with different pH values (8.3, 8.0, 7.5, 7.0, 6.5, and 6.0) for 96 h. Crack-cocaine in the different acidification scenarios triggered cyto-genotoxicity, which affected the overall health of mussels exposed to cocaine environmentally relevant concentration. This study produced the first data on biomarker responses associated with CO2-induced acidification and illicit drugs (crack-cocaine) in marine organisms.

Effects of CO2 enrichment on two microalgae species: A toxicity approach using consecutive generations.

As a result of the increasing pressure provoked by anthropogenic activities, the world climate is changing and oceans health is in danger. One of the most important factors affecting the marine environment is the well-known process called ocean acidification. Also, there are other natural or anthropogenic processes that produce an enrichment of CO2 in the marine environment (CO2 leakages from Carbon Capture and Storage technologies (CCS), organic matter diagenesis, volcanic vents, etc). Most of the studies related to acidification of the marine environment by enrichment of CO2 have been focused on short-term experiments. To evaluate the effects related to CO2 enrichment, laboratory-scale experiments were performed using the marine microalgae Tetraselmis chuii and Phaeodactylum tricornutum. Three different pH values (two treatments - pH 7.4 and 6.0 - and a control - pH 8.0) were tested on the selected species across four consecutive generations. Seawater was collected and exposed to different scenarios of CO2 enrichment by means of CO2 injection. The results showed different effects depending on the species and the generation used. Effects on T. chuii were shown on cell density, chlorophyll-a and metabolic activity, however, a slight adaptation across generations was found in this last parameter. P. tricornutum was more sensitive to acidification conditions through generations, with practically total growth inhibition in the fourth one. The conclusions obtained in this work are useful to address the potential ecological risk related to acidification by enrichment of CO2 on the marine ecosystem by using consecutive generations of microalgae.

Metal fractionation in marine sediments acidified by enrichment of CO2: A risk assessment.

Carbon-capture and storage is considered to be a potential mitigation option for climate change. However, accidental leaks of CO2 can occur, resulting in changes in ocean chemistry such as acidification and metal mobilization. Laboratory experiments were performed to provide data on the effects of CO2-related acidification on the chemical fractionation of metal(loid)s in marine-contaminated sediments using sequential extraction procedures. The results showed that sediments from Huelva estuary registered concentrations of arsenic, copper, lead, and zinc that surpass the probable biological effect level established by international protocols. Zinc had the greatest proportion in the most mobile fraction of the sediment. Metals in this fraction represent an environmental risk because they are weakly bound to sediment, and therefore more likely to migrate to the water column. Indeed, the concentration of this metal was lower in the most acidified scenarios when compared to control pH, indicating probable zinc mobilization from the sediment to the seawater.

Bioavailability and toxicity of metals from a contaminated sediment by acid mine drainage: linking exposure-response relationships of the freshwater bivalve Corbicula fluminea to contaminated sediment.

Streams and rivers strongly affected by acid mine drainage (AMD) have legal vacuum in terms of assessing the water toxicity, since the use of conventional environmental quality biomarkers is not possible due to the absence of macroinvertebrate organisms. The Asian clam Corbicula fluminea has been widely used as a biomonitor of metal contamination by AMD in freshwater systems. However, these clams are considered an invasive species in Spain and the transplantation in the field study is not allowed by the Environmental Protection Agency. To evaluate the use of the freshwater bivalve C. fluminea as a potential biomonitor for sediments contaminated by AMD, the metal bioavailability and toxicity were investigated in laboratory by exposure of clams to polluted sediments for 14 days. The studied sediments were classified as slightly contaminated with As, Cr, and Ni; moderately contaminated with Co; considerably contaminated with Pb; and heavily contaminated with Cd, Zn, and specially Cu, being reported as very toxic to Microtox. On the fourth day of the exposure, the clams exhibited an increase in concentration of Ga, Ba, Sb, and Bi (more than 100 %), followed by Co, Ni, and Pb (more than 60 %). After the fourth day, a decrease in concentration was observed for almost all metals studied except Ni. An allometric function was used to determine the relationship between the increases in metal concentration in soft tissue and the increasing bioavailable metal concentrations in sediments.

Yes, caffeine, ibuprofen, carbamazepine, novobiocin and tamoxifen have an effect on Corbicula fluminea (Müller, 1774).

Reports indicating the presence of pharmaceutical in fresh water environment in the ngL(-1) to µgL(-1) range are occurring with increasing frequency. It is also a fact that pharmaceuticals may produce adverse effects on aquatic organisms. Nevertheless, there is still a lack of knowledge regarding how these emergent contaminants may affect aquatic biota. The goal of this research was to evaluate the sublethal responses in Corbicula fluminea such as, general stress (lysosomal membrane stability [LMS]), biomarkers of phase I and II (etoxyresorufin O-deethylase [EROD], dibenzylfluorescein dealkylase [DBF], gluthathione-S-transferase [GST]), oxidative stress (gluthathione reductase [GR], gluthathione peroxidase [GPX], lipid peroxidation [LPO]), and biomarkers of effect (DNA damage) after 21 days of exposure to caffeine, ibuprofen, carbamazepine, novobiocin and tamoxifen at 0.1, 1, 5, 10, 15, 50µgL(-1). Environmental concentrations tested in this study caused general stress and produced changes on biomarkers tested. LMS, responses from phase I and II enzymatic activity, oxidative stress, and biomarker of effect represent important ecotoxicological information, and will provide a useful reference for the assessment of selected drugs and the effects which these compounds may have on aquatic invertebrates, using C. fluminea as a bioindicator species.

Effects on the mobility of metals from acidification caused by possible CO2 leakage from sub-seabed geological formations.

Carbon dioxide capture and storage (CCS) in submarine geological formations has been proposed as a mitigation measure for the prevention of global warming. However, leakage of CO2 to overlying sediments may occur over time, leading to various effects on ecosystems. Laboratory-scale experiments were performed, involving direct release of carbon dioxide into sediment, inside non-pressurized chambers, in order to provide data on the possible effects of CO2 leakage from geological storage sites on the fate of several metals. Marine sediments from three sites with different levels of contamination were sampled and submitted to acidification by means of CO2 injection. The experiment lasted 10 days and sediment samples were collected at the beginning and end of the experiment and pore water was extracted for metal analysis. The results revealed that mobility of metals from sediment to pore water depends on the site, metal and length of time exposed. Mobilization of the metals Al, Fe, Zn, Co, Pb and Cu increases with acidification, and this response generally increases with time of exposure to CO2 injection. The geochemical model applied suggests that acidification also influences the speciation of metals, transforming metals and metalloids, like As, into species much more toxic to biota. The data obtained from this study will be useful for calculating the potential risk of CCS activities to the marine environment.

Using lysosomal membrane stability of haemocytes in Ruditapes philippinarum as a biomarker of cellular stress to assess contamination by caffeine, ibuprofen, carbamazepine and novobiocin.

Although pharmaceuticals have been detected in the environment only in the range from ng/L to microg/L, it has been demonstrated that they can adversely affect the health status of aquatic organisms. Lysosomal membrane stability (LMS) has previously been applied as an indicator of cellular well-being to determine health status in bivalve mussels. The objective of this study is to evaluate LMS in Ruditapes philippinarum haemolymph using the neutral red retention assay (NRRA). Clams were exposed in laboratory conditions to caffeine (0.1, 5, 15, 50 microg/L), ibuprofen (0.1, 5, 10, 50 microg/L), carbamazepine and novobiocin (both at 0.1, 1, 10, 50 microg/L) for 35 days. Results show a dose-dependent effect of the pharmaceuticals. The neutral red retention time measured at the end of the bioassay was significantly reduced by 50% after exposure to environmental concentrations (p < 0.05) (caffeine = 15 microg/L; ibuprofen = 10 microg/L; carbamazepine = 1 microg/L and novobiocin = 1 microg/L), compared to controls. Clams exposed to these pharmaceuticals were considered to present a diminished health status (retention time < 45 min), significantly worse than controls (96 min) (p < 0.05). The predicted no environmental effect concentration (PNEC) results showed that these pharmaceuticals are very toxic at the environmental concentrations tested. Measurement of the alteration of LMS has been found to be a sensitive technique that enables evaluation of the health status of clams after exposure to pharmaceuticals under laboratory conditions, thus representing a robust Tier-1 screening biomarker.

Bioaccumulation and effects of metals bound to sediments collected from Gulf of Cádiz (SW Spain) using the polychaete Arenicola marina.

A short-term whole-sediment test using the polychaete Arenicola marina was conducted under laboratory conditions to assess the bioavailability of metals bound to sediments collected from 12 sites of the Gulf of Cádiz. To achieve this objective, the rate of increase of metal bioaccumulation and the induction of a typical biomarker, metallothioneinlike proteins (MTLPs) were determined. Results of the multivariate analysis showed associated metal-rich sediments, increased rate of Cu and Zn accumulations, but lower toxicity with an increased MTLP induction, whereas sedimentary Ni and Co concentrations were related to higher toxicity to lugworms, although it might be caused by other contaminants present in these sediments. The linear kinetic approach was shown to be valid in certain circumstances, but more validation studies of this parameter are required before it can be recommended for use in evaluating metal bioavailability in sediments.

Toxicity and potential risk assessment of a river polluted by acid mine drainage in the Iberian Pyrite Belt (SW Spain).

Metal contamination from acid mine drainage (AMD) is a serious problem in the southwest of the Iberian Peninsula, where the Iberian Pyrite Belt is located. This zone contains original sulfide reserves of about 1700Mt distributed among more than 50 massive sulfide deposits. Weathering of these minerals releases to the waters significant quantities of toxic elements, which severely affect the sediments and surface waters of the region. The main goal of this paper is to evaluate the toxicity and the potential risk associated with the mining areas using Microtox test and different factors which assess the degree of contamination of the sediments and waters. For this, a natural stream polluted by AMD-discharge from an abandoned mine has been studied. The results show that elevated concentrations of Cu, As and Zn involve an important potential risk on the aquatic environment, associated both with sediments and waters. Microtox test informs that the sediments are extremely or very toxic, mainly related to concentrations of Fe, As, Cr, Al, Cd, Cu and Zn. Pollution is mainly transferred to the sediments increasing their potential toxicity. A natural creek affected by AMD can store a huge amount of pollution in its sediments while exhibiting a not very low water pH and low water metal concentration.

Ecological risk assessment of sediment management areas: application to Sado Estuary, Portugal.

The purpose of this work was to integrate different methodologies to assess the potential ecological risk of estuarine sedimentary management areas, using the Sado Estuary in Portugal as case study. To evaluate the environmental risk of sediment contamination, an integrative and innovative approach was used involving assessment of sediment chemistry, sediment toxicity, benthic community structure, human driving forces and pressures and management areas organic load levels. The basis for decision-making for overall assessment was a statistical multivariate analysis appended into a score matrix tables, using a best expert judgment. The integrated approach allowed to identify from the 19 management areas analyzed, three with no risk but other three with high risk to cause adverse effects in the biota, related with the contaminants analyzed. The methodologies used showed to be effective as a support for decision making leading to future estuarine management recommendations.

Toxicity and bioaccumulation of copper and lead in five marine microalgae.

On five marine microalgae with the same biovolume quantity (Tetraselmis chuii, Rhodomonas salina, Chaetoceros sp., Isochrysis galbana (T-iso) and Nannochloropsis gaditana) 72-h exposure toxicity tests with copper and lead were performed. For both metals, 72-h EC50s showed T. chuii as the most tolerant and R. salina as one of the most sensitive. Besides copper and lead EC(50) concentrations, metal concentrations in solution and accumulated on/in the cell where also analysed. T. chuii, the most tolerant species accumulated high copper concentrations (EC(50(Cu))=330 microgL(-1); EC(50(Pb))=2600 microgL(-)1), and R. salina the most sensitive to copper, accumulated the highest amount of this metal (EC(50(Cu))=50 microgL(-1)). Results of this study show that there is no specific relationship between cell tolerance and accumulated metal on/in the cell. On the other hand, due to an established evidence of the influence of cellular density in microalgae toxicity tests, this effect was also studied. Results showed reduced EC(50) values when initial cellular densities decreased. In this study, the term "toxic cellular quota" was used to express all data. This allowed, in a single expression, the combination of two parameters that clearly influence growth, cellular density and toxic concentration.

Toxicokinetic approach for the assessment of endocrine disruption effects of contaminated dredged material using female Carcinus maenas.

Ecotoxicological effects associated with contaminants present in dredged material from three Spanish ports, Ría of Huelva (SW, Spain), La Coruña (NW, Spain) and Bay of Cádiz (SW, Spain) were determined using a marine biotest based on endocrine disruption effects (vitellogenesis process alteration). Intermoult female Carcinus maenas were exposed in the laboratory to sediments from the Spanish ports per replicate during 21 days. Crab haemolymph samples were taken for vitellogenin/vitellin (VTG) analysis on days 0, 7, 14 and 21. Furthermore, chemical analysis was performed in the stations to determine the degree and nature of sediment contamination (Cr, Ni, Cu, Zn, Cd, Pb, Hg, As, PAHs and PCBs). A significant induction (P < 0.05) in vitellogenin/vitellin concentration was observed over time in individuals exposed to sediment samples containing significant concentrations of PAHs, PCBs, Hg, Pb and Zn. The toxicokinetic approach, proposed in this study related to the use of this biomarker in C. maenas, proved to be a powerful and sensitive tool to evaluate toxicity effects associated with contaminants present in dredged material. Moreover the integration of the results obtained through multivariate analysis approach (MAA) allowed the identification contaminants bound to sediments associated with adverse effects, validating the use of this marine biotest in a regulatory framework.

The use of a metallothionein-like-proteins (MTLP) kinetic approach for metal bioavailability monitoring in dredged material.

Ecotoxicological effects associated with contaminants present in dredged material from Spain were determined using a marine biotest based on the determination of metallothionein-like-protein concentrations (MTLPs) in the shore crab Carcinus maenas. Intermoult female C. maenas were exposed in the laboratory to sediments from three Spanish ports, Ría de Huelva (SW, Spain), La Coruña (NW, Spain) and Bahía de Cádiz (SW, Spain) per replicate during 21 days. Hepatopancreas samples from crabs were taken for metallothioneins analysis on days 0, 7 and 21. Furthermore, chemical analysis was performed in the stations to determine the degree and nature of sediment contamination (Cr, Ni, Cu, Zn, Cd, Pb, Hg, As, PAHs and PCBs). A significant increase (p<0.05) in metallothionein concentration was observed over time in individuals exposed to sediment from the port Ría de Huelva characterized by high concentrations of metals. A toxicokinetic approach is proposed in this study related to the use of this biomarker in C. maenas to evaluate bioavailability associated with metals present in dredged material. As a first step, this toxicokinetic approach might reveal as a sensitive tool for evaluating bioavailability of contaminants present in dredged material.

Biological adverse effects on bivalves associated with trace metals under estuarine environments.

Toxic effects of pollutants on marine organisms can be studied both by performing field measurements, and by undertaking laboratory simulation experiments. Here is described the effect of trace metals Zn, Cd, Pb and Cu on the clam Scrobicularia plana along a salinity gradient simulated in a hypothetical estuary using simulation experiments. The simulator produces a continuous entry of trace metals into the estuary through injection in the lower salinity tank of the system. The clams were exposed during two weeks to different concentration of trace metals to assess the bioaccumulation process along a salinity gradient. Bivalves were analysed for body tissue residue to determine the bioaccumulation factors related to each metal and the salinity influence was addressed. Differences among tanks were observed as a result of the salinity gradient. In the achieved assays, the mechanism of bioaccumulation of Zn and Cd in organisms was more efficient at high salinity values. Bioaccumulation factors for both metals showed a linear increase with the increase of salinity values. It seems that the mechanism of bioaccumulation of Pb and Cu in organisms was dependent on two simultaneous processes: the proximity to the input point of metals and the low salinity values.

Sediment quality in the Atlantic coast of Spain.

Sediments from the Atlantic coast of Spain have been studied to evaluate environmental quality by using an integrated approach including chemical and toxicological data. Sediment samples were collected in four littoral ecosystems located in Spain, Bay of Cádiz, Guadalquivir River estuary, Ría of Huelva, and Ría of Coruña. To characterize the sediments, organic carbon, granulometric content, total sulfide, eight trace metals (Hg, Cd, Pb, Cu, Zn, As, Ni, and Cr), polychlorinated biphenyls (PCBs), and polycyclic aromatic hydrocarbons (PAHs) were measured. The toxicity of sediments was assessed with the amphipod Ampelisca brevicornis, the clam Ruditapes philippinarum, juveniles of the fish Solea senegalensis, populations of the estuarine rotifer Brachionus plicatilis, and populations of the bacterium Vibrio fischeri (Microtox). The results obtained show that in general, stations located in the Ría of Huelva were associated with heavy metal contamination and with the highest toxicity. Only chronic toxicity tests were capable of identifying the effects associated with PCB concentrations. The sediment quality guidelines calculated by means of a multivariate analysis approach for contaminants not associated with biological effects (mg/kg) are Hg, 0.54; Cd, 0.51; Pb, 260; Cu, 209; Zn, 513; As, 27.4; and total PCBs, 0.05.