Cátia Velez

Looking for suitable biomarkers in benthic macroinvertebrates inhabiting coastal areas with low metal contamination: Comparison between the bivalve Cerastoderma edule and the Polychaete Diopatra neapolitana

Metals accumulated in marine sediments are often a threat to benthic communities. With the recognized importance and wide use of stress biochemical responses as indicators of metal contamination it becomes essential to compare these markers between different species and verify their ubiquity and accuracy. Using wild Diopatra neapolitana and Cerastoderma edule, collected at several areas differing in metal contamination, this study aimed to assess the use of these two macrobenthic species as sentinel organisms and to determine the applicability of currently used biomarkers in benthic species exposed to a range of low metal and As concentrations. Total metal accumulation and intracellular partitioning was analyzed and metal-induced alterations were assessed through the analysis of several biochemical parameters in both organisms, including stress-induced reactive oxygen species (ROS), lipid peroxidation and protein content, the activity of antioxidant enzymes catalase (CAT), superoxide dismutase (SOD) and of glutathione S-transferase (GST). Metal chelation by Metallothioneins (MTs) was also determined. Results revealed that D. neapolitana accumulated higher levels of metals when compared to C. edule, independently of the sediment concentration. Results also showed strong species-specific responses to metals and differences in the ability to sequester metals. Overall, C. edule showed to be more efficient metal chelator and precipitatior than D. neapolitana, which was less tolerant and presented oxidative stress. MTs proved to be a good predictor of metal accumulation in both species, even under low metal exposures. On the other hand, lipid peroxidation was a good indicator of oxidative damage, only observed in D. neapolitana, which was a result of higher metal retention in the soluble fraction.

Physiological and biochemical responses of three Veneridae clams exposed to salinity changes.

Given their global importance, coastal marine environments are a major focus of concern regarding the potential impacts of climate change, namely due to alterations in seawater salinity. It is known that environmental characteristics, such as salinity, affect immune and physiological parameters of bivalves. Nevertheless, scarce information is available concerning the biochemical alterations associated with salinity changes. For this reason, the present work aimed to evaluate the biochemical responses of three venerid clam species (Venerupis decussata, Venerupis corrugata, Venerupis philippinarum) submitted to salinity changes. The effects on the native (V. decussata and V. corrugata) and invasive (V. philippinarum) species collected from the same sampling site and submitted to the same salinity gradient (0 to 42g/L) were compared. The results obtained demonstrated that V. corrugata is the most sensitive species to salinity changes and V. decussata is the species that can tolerate a wider range of salinities. Furthermore, our work showed that clams under salinity associated stress can alter their biochemical mechanisms, such as increasing their antioxidant defenses, to cope with the higher oxidative stress resulting from hypo and hypersaline conditions. Among the physiological and biochemical parameters analyzed (glycogen and protein content; lipid peroxidation levels, antioxidant enzymes activity; total, reduced and oxidized glutathione) Catalase (CAT) and especially superoxide dismutase (SOD) showed to be useful biomarkers to assess salinity impacts in clams.

Physiological and biochemical responses of the Polychaete Diopatra neapolitana to organic matter enrichment

Several studies have demonstrated that organic matter enrichment may be associated to aquaculture, leading to impoverished benthic communities and species succession with loss of biodiversity, but very few studies have investigated biochemical and physiological alterations that species affected by aquaculture activities undergo. Thus, in the present study, the effects of the organic enrichment originating from an oyster culture were studied in the Polychaete Diopatra neapolitana, a species already shown to be sensitive to inorganic contamination. For this, physiological responses and biochemical alterations were evaluated. The results obtained revealed that individuals from highly organically enriched areas presented lower capacity to regenerate their body but higher glycogen and protein levels. Furthermore, with increasing organic matter D. neapolitana increased the lipid peroxidation (LPO), the oxidized glutathione content (GSSG) and Glutathione S-transferase activity (GSTs) content, and the activity of the antioxidant enzymes catalase (CAT) and superoxide dismutase (SOD). This study evidenced that organic matter enrichment induced biochemical and physiological alterations in D. neapolitana. Thus, this species was shown to be a good sentinel species to monitor organic contamination.

The effects of carbamazepine on macroinvertebrate species: Comparing bivalves and polychaetes biochemical responses.

In the present study, the bivalve Scrobicularia plana and the polychaete Diopatra neapolitana were exposed to an increasing carbamazepine (CBZ) concentration gradient. Both species are among the most widely used bioindicators, and CBZ is one of the most commonly found drugs in the aquatic environment. After a chronic exposure (28 days), the results obtained revealed that CBZ induced biochemical alterations in both species. Our findings demonstrated that S. plana and D. neapolitana reduced the CBZ accumulation rate at higher CBZ concentrations, probably due to their capacity to decrease their feeding rates at stressful conditions. Nevertheless, this defence mechanism was not enough to prevent both species from oxidative stress. In fact, S. plana and D. neapolitana were not able to efficiently activate their antioxidant defence mechanisms which resulted in the increase of lipid peroxidation, especially at the highest CBZ concentrations. Comparing both species, it seems that S. plana was the most sensitive species since stronger biochemical alterations were observed in this species.

The impacts of pharmaceutical drugs under ocean acidification: New data on single and combined long-term effects of carbamazepine on Scrobicularia plana

Ocean acidification and increasing discharges of pharmaceutical contaminants into aquatic systems are among key and/or emerging drivers of environmental change affecting marine ecosystems. A growing body of evidence demonstrates that ocean acidification can have direct and indirect impacts on marine organisms although combined effects with other stressors, namely with pharmaceuticals, have received very little attention to date. The present study aimed to evaluate the impacts of the pharmaceutical drug Carbamazepine and pH 7.1, acting alone and in combination, on the clam Scrobicularia plana. For this, a long-term exposure (28 days)was conducted and a set of oxidative stress markers was investigated. The results obtained showed that S. plana was able to develop mechanisms to prevent oxidative damage when under low pH for a long period, presenting higher survival when exposed to this stressor compared to CBZ or the combination of CBZ with pH 7.1. Furthermore, the toxicity of CBZ on S. plana was synergistically increased under ocean acidification conditions (CBZ + pH 7.1): specimens survival was reduced and oxidative stress was enhanced when compared to single exposures. These findings add to the growing body of evidence that ocean acidification will act to increase the toxicity of CBZ to marine organisms,which has clear implications for coastal benthic ecosystems suffering chronic pollution from pharmaceutical drugs.

Ruditapes philippinarum and Ruditapes decussatus under Hg environmental contamination

The native species Ruditapes decussatus and the invasive species Ruditapes philippinarum have an important ecological role and socio-economic value, from the Atlantic and Mediterranean to the Indo-Pacific region. In the aquatic environment, they are subjected to the presence of different contaminants, such as mercury (Hg) and its methylated form, methylmercury (MeHg). However, few studies have assessed the impacts of Hg on bivalves under environmental conditions, and little is known on bivalve oxidative stress patterns due to Hg contamination. Therefore, this study aims to assess the Hg contamination in sediments as well as the concentration of Hg and MeHg in R. decussatus and R. philippinarum, and to identify the detoxification strategies of both species living in sympatry, in an aquatic system with historical Hg contamination. The risk to human health due to the consumption of clams was also evaluated. The results obtained demonstrated that total Hg concentration found in sediments from the most contaminated area was higher than the maximum levels established by Sediment Quality Guidelines. This study further revealed that the total Hg and MeHg accumulation in both species was strongly correlated with the total Hg contamination of the sediments. Nonetheless, the THg concentration in both species was lower than maximum permissible limits (MPLs) of THg defined by international organizations. R. decussatus and R. philippinarum showed an increase in lipid peroxidation levels along with the increase of THg accumulation by clams. Nevertheless, for both species, no clear trend was obtained regarding the activity of antioxidant (superoxide dismutase, catalase) and biotransformation (glutathione S-transferase) enzymes and metallothioneins with the increase of THg in clams. Overall, the present work demonstrated that both species can be used as sentinel species of contamination and that the consumption of these clams does not constitute a risk for human health.

Native and introduced clams biochemical responses to salinity and pH changes

By the end of year 2100 physiological and biochemical performance of aquatic organisms are expected to become strongly affected by salinity and pH shifts, which in turn may favor the conditions for introduced species to invade new ecosystem areas. Given this, we evaluated the effects of salinity and pH changes in native Ruditapes decussatus and introduced Ruditapes philippinarum clams, by measuring different biomarkers related to oxidative stress, metabolic activity and osmoregulation capacity. Results showed that extreme salinities induced mortality in both species, while all clams survived under low pH (7.3). Both species mobilized glycogen as a source of energy towards cells protection mechanisms under extreme salinities. The native species presented higher lipid peroxidation levels while the introduced species was able to prevent oxidative damages through the induction of antioxidant enzymes at most extreme salinities. R. philippinarum also induced CA activity to balance the ion homeostasis at extreme salinities. In contrast, low pH induced oxidative damages, an increase of antioxidant (catalase), detoxification (glutathione S-transferases) and osmoregulation (carbonic anhydrase) mechanisms in R. philippinarum compared to the native clams. Overall, salinity and pH changes can alter physiological and biochemical status of native and introduced clam species.

Combined effects of seawater acidification and salinity changes in Ruditapes philippinarum

Due to human activities, predictions for the coming years indicate increasing frequency and intensity of extreme weather events (rainy and drought periods) and pollution levels, leading to salinity shifts and ocean acidification. Therefore, several authors have assessed the effects of seawater salinity shifts and pH decrease on marine bivalves, but most of these studies evaluated the impacts of both factors independently. Since pH and salinity may act together in the environment, and their impacts may differ from their effects when acting alone, there is an urgent need to increase our knowledge when these environmental changes act in combination. Thus, the present study assessed the effects of seawater acidification and salinity changes, both acting alone and in combination, on the physiological (condition index, Na and K concentrations) and biochemical (oxidative stress related biomarkers) performance of Ruditapes philippinarum. For that, specimens of R. philippinarum were exposed for 28days to the combination of different pH levels (7.8 and 7.3) and salinities (14, 28 and 35). The results obtained showed that under control pH (7.8) and low salinity (14) the physiological status and biochemical performance of clams was negatively affected, revealing oxidative stress. However, under the same pH and at salinities 28 and 35 clams were able to maintain/regulate their physiological status and biochemical performance. Moreover, our findings showed that clams under low pH (7.3) and different salinities were able to maintain their physiological status and biochemical performance, suggesting that the low pH tested may mask the negative effects of salinity. Our results further demonstrated that, in general, at each salinity, similar physiological and biochemical responses were found in clams under both tested pH levels. Also, individuals under low pH (salinities 14, 28 and 25) and exposed to pH 7.8 and salinity 28 (control) tend to present a similar response pattern. These results indicate that pH may have a lower impact on clams than salinity. Thus, our findings point out that the predicted increase of CO2 in seawater and consequent seawater acidification will have fewer impacts on physiological and biochemical performance of R. philippinarum clams than salinity shifts.

Biochemical performance of native and introduced clam species living in sympatry: The role of elements accumulation and partitioning

The present study reports metal and arsenic contamination in sediments, as well as element accumulation and partitioning in native (Ruditapes decussatus and Venerupis corrugata) and introduced (Ruditapes philippinarum) clam species living in sympatry at the Óbidos lagoon (Portugal). The biochemical performance and the human health risks derived from the consumption of these species are also discussed. The results obtained showed that R. decussatus was the most abundant species in all the sampling sites, revealing that the introduced clam has not yet supplanted the native species. The concentration of elements was higher in areas with higher Total Organic Matter (TOM) and fines content, being Chromium (Cr), Copper (Cu) and Lead (Pb) the most abundant metals. Clams from these areas showed the highest concentration of elements but the lowest bioaccumulation levels. Furthermore, except for As, higher concentration of elements was found in clams insoluble fraction, the less toxic fraction to the organisms. Due to the low contamination levels and because elements, except As, were mainly allocated to the insoluble fraction, clams presented similar biochemical parameters among distinct areas, with no significant oxidative stress induced. Furthermore, clams from the Óbidos lagoon represent a low health risk to human consumption since, except for As, their contamination levels were below the maximum permissible limits defined by international organizations.

The impacts of As accumulation under different pH levels: Comparing Ruditapes decussatus and Ruditapes philippinarum biochemical performance.

Marine bivalves have been used to assess environmental As contamination and the effects of seawater acidification when both factors are acting alone, but limited information is available regarding the impacts of both factors acting in combination. The aim of this study was to compare physiological (glycogen) and biochemical (lipid peroxidation, superoxide dismutase, catalase, glutathione-S-transferase and alkaline phosphatase) responses in both native (Ruditapes decussatus) and introduced (R. philippinarum) clams, when exposed to the combined effects of pH (7.8, control; 7.3) and As concentrations (0 and 4mg/L). The combined effect of As and pH on the health risks associated with clam consumption was also analyzed. Results revealed that both species were able to accumulate As under both pH levels, although higher As concentrations where observed under low pH. Thus, predicted pH decrease will potentiate health risks associated with the consumption of such species, since less amount of clams exposed to As is needed for an adult to exceed the provisional tolerable weekly intake (PTWI). Low pH, As exposure and the combination of both factors did not negatively affect the native species, since clams were able to maintain their physiological and biochemical performance among all conditions. On the other hand, R. philippinarum was negatively affected by As exposure at control pH (7.8), inducing biotransformation and antioxidant defense mechanisms against As toxicity. R. philippinarum exposed and non-exposed to As presented similar responses under low pH although at this condition the introduced species accumulated twice the amount of As than R. decussatus.