Application of bioassays in toxicological hazard, risk and impact assessments of dredged sediments

Given the potential environmental consequences of dumped dredged harbour sediments it is vital to establish the potential risks from exposure before disposal at sea. Currently, European legislation for disposal of contaminated sediments at sea is based on chemical analysis of a limited number of well-known contaminants for which maximum acceptable concentrations, action levels (ALs), have been set. The present paper addresses the issue of the applicability of in vitro and in vivo bioassays for hazard, risk and local impact assessment of dredged polluted sediments to be disposed of at sea. It discusses how and to what extent selected bioassays can fill in the gaps left open by chemical analysis and the way in which the bioassays may contribute to the present licensing system for disposal. Three different purposes for application were distinguished: the most basic application (A) is a rapid determination of the hazard (potential toxicity) of dredged sediments which is then compared to ALs in a licensing system. As with chemical analysis on whole sediment extracts, the bioavailability of the chemicals is not taken into account. As in vitro assays with sediment extracts are not sensitive to matrix effects, a selection of specific in vitro bioassays can be suitable fast and standardized additions for the licensing system. When the outcome of (A) does not convincingly demonstrate whether the sediment is clean enough or too polluted, further bioanalysis can help the decision making process (B). More aspects of the mostly unknown complex chemical mixtures are taken into account, including the bioavailability and chronic toxicity focusing on ecologically relevant endpoints. The ecotoxicological pressure imposed by the dredged sediments can be quantified as the potentially affected fraction (PAF) based on chemical or biological analysis of levels of contaminants in sediment or biota. To validate the predicted risk, the actual impact of dumped harbour sediments on local ecosystems (C) can be determined using a dedicated set of in vitro and in vivo bioassays as well as bio-indicators selected based on the information obtained from (A) and (B) and on the characteristics of the local ecosystem. Conversely, the local sediment impact assessment (C) can direct fine-tuning of the selection of chemical and bioassay analyses and for setting safe levels in the licensing system. It is concluded that in vitro and in vivo bioassays and biological indicators are useful tools in the process of hazard, ecotoxicological risk and impact assessment of dredged harbour sediments, provided they are consciously chosen and quality criteria for assay performance are defined.     

Integrative assessment of coastal pollution: Development and evaluation of sediment quality criteria from chemical contamination and ecotoxicological data

Elutriate embryo-larval bioassays with sea-urchins (Paracentrotus lividus) were conducted concurrently with chemical analyses of sediments and biota as part of an integrative assessment of pollution in highly productive coastal regions. High metal contents and organic compounds in sediments and mussels were found in localised areas from the inner part of the estuaries indicating a clear anthropogenic influence. In particular, average maximum concentrations of 2803 mg Cu/kg dw, 776 mg Pb/kg dw, 2.5 mg Hg/kg dw and 5803 μg ∑₇PAHs/kg dw were measured in sediments from the most polluted sites. Significant correlations were observed between sediment chemistry and toxicity bioassays. Moreover, the Mantel test revealed a significant correlation (r_M=0.80; p<0.01) between sediment pollutant concentrations and toxicity data profiles. In addition, sediment quality criteria were used to help in the ecological interpretation of sediment chemistry data and to identify pollutants of concern. The toxicity bioassays identified polluted sites and quantified the level of toxicity, providing a cost-effective tool to complement the routine chemical monitoring currently conducted in European coastal waters with ecologically relevant information. This is in line with the recent European legislation that advocates the use of biological tools with the ultimate aim of protecting marine resources from anthropogenic substances that will affect their sensitive early life stages.     

Toxicity evaluation by using intact sediments and sediment extracts

The toxicity of intact sediments and sediment extracts, from both an uncontaminated site and a site contaminated by pulp-mill effluents, was tested in a five months study. The deposit-feeding amphipod Monoporeia affinis was exposed in soft-bottom flow-through water microcosms. To examine potential toxicity a set of reproduction endpoints was used including fecundity and different embryo aberrations such as malformed eggs. Among extracts, the aliphatic/monoaromatic and diaromatic fractions along with the total extract were shown to cause the highest toxicity measured as malformed eggs, while the polyaromatic fraction caused toxicity at background levels. A comparison between sediment extracts and pulp mill contaminated intact sediment, however, showed no toxicity of the intact sediment. Thus, the extraction procedure seems to increase bioavailability and subsequently toxicity as compared to the intact sediments in situ. In toxicity testing using fractionated extracts of sediments in a toxicity identification evaluation (TIE) procedures, caution should therefore be taken when assessing bioavailable contaminants in contaminated areas. This should be taken in account both in determining remediation priorities as well as in ecological risk assessments.     

Effects of new antifouling compounds on the development of sea urchin

Tributyltin oxide (TBTO) has been used worldwide in marine antifouling paints as a biocide for some time. However, it produced toxic effects, especially in marine water/sediment ecosystems. Consequently, its use in antifouling paints has been prohibited in many countries. In this study, the toxicity of alternative and/or new antifouling biocides compared with TBTO is assessed by a biological method. The effects of these chemicals on marine species have not been well studied. This paper assesses, comparatively, the effects of eight biocides on sea urchin eggs and embryos. The chemicals assessed were TBTO, Irgarol 1051, M1 (the persistent degradation product of Irgarol), Diuron, zinc pyrithione, 'KH101', 'Sea-Nine 211', and copper pyrithione. For these chemicals, toxicity appears to be in the order zinc pyrithione > Sea-Nine 211 > KH101 > copper pyrithione > TBTO > Diuron approximately = Irgarol 1051 > M1. Here, we show that zinc pyrithione, Sea-Nine 211, KH101, and copper pyrithione are much more toxic to sea urchins than TBTO or the other chemicals.     

Toxicity Assessment by Microtox® in Sediments,Pore Waters and Sediment Saline Elutriates in the Gulf of Gdansk (Baltic Sea)

The toxicity of sediments in the Gulf of Gdansk is analyzed in relation to the chemical composition of interstitial and near‐bottom waters, and sediment properties. The toxicity of sediments, pore waters and saline elutriates is determined by using the Microtox® test based on changes in light production of the luminescent bacteria Vibrio fischeri. The results indicate considerable toxicity in the majority of examined sediments. Since the sediment elutriates and pore waters are toxic in some cases, the total toxicity of the sediments is likely to be due to both sediment‐bound and water soluble substances. The sediment toxicity is related to the percentage contribution of the fine fraction of sediments. A significant correlation between the toxicity of the sediments and the black carbon content implies anthropogenic contamination. The toxicity of the sediments is seen to increase with the increase of hydrogen sulfide concentration in pore waters. The ammonia in pore waters was found not to be responsible for the toxicity of the sediments.     

Burrowing responses of the short-neck clam Ruditapes philippinarum to sediment contaminants

The burrowing responses of a common tropical bivalve, the short-neck clam Ruditapes philippinarum, to cadmium (Cd)-spiked sediment, variations of sediment grain size and natural sediments collected from 15 locations in Hong Kong's inshore waters were investigated through a series of laboratory tests. Results showed that the burrowing response exhibited a negative relationship with an increase in Cd concentration in the spiked sediments. The level of Cd was also found to be directly proportional to the percent mortality of the clam. However, there was no significant difference (p>0.05) in the time elapsed for the clam to burrow into sediments with different grain size composition. The elapsed time for 50% of the test clams to burrow into the sediment (ET₅₀) over a period of 48 h was calculated for the sediment samples collected from the 15 locations. Results of ANOVA showed significant difference (p<0.05) among the sediment samples. Tukey's multiple comparison test revealed two groups of sediments: group 1 containing 3 sediment samples collected from Victoria Harbour and group 2 containing 12 samples obtained from other coastal areas of Hong Kong. The ET₅₀ value for group 1 sediments was found to be greater than 2880 min whilst the ET₅₀ for group 2 sediments had a mean of 173.9 min. Agglomerative classification of the sediment samples, based on metal content (As, Cd, Cr, Cu, Ni, Pb, Zn), also showed two similar groups, suggesting that the ET₅₀ values were correlated with the metal level of the sediment samples. Group 1 sediments collected from Victoria Harbour had greater concentrations of Cd, Cr, Cu, Ni and Zn as compared to that in group 2. The present study demonstrated that high metal content in the sediment tends to inhibit the burrowing responses of the clam and that there is potential to develop the burrowing responses of R. philippinarum as a sublethal sediment toxicity test.     

A toxicity identification evaluation of silty marine harbor sediments to characterize persistent and non-persistent constituents

Sediment toxicity in silty marine harbor sediments is frequently dominated by ammonia or sulfide, leaving the adverse effects of persistent toxic substances unnoticed. To investigate the latter, we subjected interstitial water from three contaminated silty sediments to toxicity identification evaluation (TIE) phase I manipulations and tested for toxicity with four bioassays: the amphipod Corophium volutator (survival as an endpoint), the sea urchin Psammechinus miliaris (fertilization, embryo development) and the bacterium Vibrio fischeri (bioluminescence inhibition).The graduated pH manipulations identified the prominent toxicity of ammonia in the amphipod and sea urchin embryo tests, and also sulfide toxicity in the bacterium test. In two of the three samples tested with the amphipods, sea urchin embryos and bacteria, a small but significant reduction in interstitial water toxicity was achieved by removing persistent compounds through C(18) solid phase extraction. EDTA chelation resulted in a slight detoxification of the interstitial water for the amphipods and sea urchin embryos, but this was not related to any measured trace metals. Despite the presence of toxic levels of ammonia and sulfide in the harbor sediments, we established the adverse biological effects of persistent constituents by means of the TIE manipulations and in vivo interstitial water bioassays.     

The toxicity of marine sediments in Victoria Harbour, Hong Kong

When the toxicity of marine sediment in Hong Kong was evaluated, it was found that the seven sediments collected within Victoria Harbour were severely contaminated with heavy metals, at concentrations many times higher than those in sediments collected from outside the harbour. The highest metal content was recorded in site VS14 (located near the airport runway and the industrialized area), with copper, zinc, lead and chromium values of 3789, 610, 138 and 601 mg kg⁻¹ dry wt, respectively. This site also had the greatest alkaline phosphatase activities (15 fluorescent intensity unit g⁻¹ wet wt), the largest number of total coliforms (910 CFU g⁻¹ wet wt) and sulphate-reducing bacteria (8.5 × 10⁴ cells g⁻¹ wet wt), implying that site VS14 was also contaminated with organic matter and nutrients. Sediment bioassays, Microtox and algal tests, demonstrated that sediment elutriates obtained from site VS14 were of greatest toxicity. The EC₁₀ value in Microtox tests was 17% elutriate, and the 96-h IC₅₀ values using Skeletonema costatum and Dunaliella tertiolecta were 40 and 79% elutriate, respectively. No toxic effects were found in sediment samples collected from the control site outside Victoria Harbour. Significant correlations were found between the results of the algal toxicity test (using S. costatum) and the coliform count and metal content of the sediments. The Microtox test was less sensitive than the algal bioassay, and no sediment elutriate, even from the site mostly contaminated by heavy metals, caused more than 50% inhibition of the light-emitting activity of the bacteria. In this study, S. costatum (the diatom) provided a more sensitive and reliable test species than D. tertiolecta (the flagellate) in differentiating the toxicity of marine sediments.     

Seasonal variation in the toxicity of sediment-associated contaminants in Corpus Christi Bay, TX

Seasonal variation in pore water and sediment-water interface (SWI) toxicity at two sites of suspected contamination was investigated using sea urchin embryological development (Arbacia punctulata) and copepod hatching success (Schizopera knabeni). Site S1, located inside a marina, was fine-grained and S2, located near a neighboring stormwater outfall, was sandy. Both sites were cored in summer and winter, along with reference sites of equivalent grain sizes. Whereas the muddy contaminated site displayed a slight decrease in sea urchin toxicity from summer to winter, pore water from the sandy contaminated site exhibited an increase in sea urchin toxicity. The sandy sites displayed slight toxicity increase to copepod hatching success in winter, which occurred in the SWI of S2 and in the pore water from the reference site, R2, where it coincided with sedimentation. Cu was the most prevalent metal in the sediment samples. Although Cu concentration increased in the whole sediments and pore waters from summer to winter, its concentration in overlying SWI exposure decreased from summer to winter. Organic compounds exhibited opposite behavior, with smaller variety identified in the winter pore water samples, but more chemicals detected in the SWI samples in winter.     

Influence of bedding orientation on the anisotropy of hydraulic conductivity in a well-sorted fluvial sediment

The paper describes a permeameter test method for determination of the hydraulic conductivity(AT) along multi-directions in fluvial sediments with cross beddings.Unlike existing in-situ permeameter methods that determine hydraulic conductivity for submerged streambeds,our method was intended to measure hydraulic conductivity of exposed streambeds or fluvial sediments.The method was applied to the Wei River,Shaanxi Province,Central China for characterization of the anisotropy of K in a well-sorted fluvial sediment.The results illustrated that even in well-sorted sediments,cross-bedding and sediment fabrication(or texture) can lead to varied K values along different measurement directions.The K value was the largest along the dip direction(or the major direction) that is parallel to the orientation of cross bedding and the smallest in the direction perpendicular to the bedding(or the minor direction). The K value in a given direction between the major and minor direction often fell in the range bounded by the K values in the major and minor directions.The anisotropy ratio of K(the ratio of K value between the major and minor directions) in two trenches for this well-sorted fluvial sediment was up to 1.14 to 1.23,respectively.Our results also demonstrated that even for well-sorted sediments,the K values between two sampling points only about 10 cm apart can differ.It is clear that the K distribution strongly correlates to the bedding orientation.     

Assessing metal toxicity in sediments using the equilibrium partitioning model and empirical sediment quality guidelines: A case study in the nearshore zone of the Bohai Sea,China

Surface sediments were collected from five nearshore (wastewater discharges, aquaculture facilities and a seaport) sites in Bohai Bay and Laizhou Bay, China. The equilibrium partitioning (EqP) model and empirical sediment quality guidelines (SQGs) were applied to assess the potential metal toxicity in the collected sediments. The results show that, based on the EqP model, 35% of stations exhibited potential metal toxicity. Several metals (Cu, Ni and Cr) exceeded the empirical SQGs (9–93% of the time), however these guidelines may not be suitable for use in the Bohai Sea owing to the background concentrations. The EqP model is a more useful method for assessing potential metal toxicity in Bohai Sea sediment than the empirical SQGs. Additionally, we have provided new understanding about methods for assessing sediment metal toxicity in the Bohai Sea that may be useful in other coastal areas in China.     

Bacterial and organic matter distribution in the sediments of the Thracian Sea (NE Aegean Sea)

Recently, black carbon has been introduced as the form of carbon that may be separated from the biologically mediated carbon cycle thereby representing the non-bioavailable fraction of the estimated organic carbon. It has been speculated that the bioavailability of organic matter may be a limiting factor for the presence of active bacteria within the sediments. In order to address this question, marine sediments were collected from the Thracian Sea (Eastern Mediterranean), a complex system impacted by riverine inputs and Black Sea water masses. In addition to counts of total bacteria, we estimated the fraction of active bacteria by using a destaining step to the DAPI staining method. Black carbon was also estimated following the thermal oxidation method in order to determine the fraction of the refractory organic matter. The fraction of black carbon to total organic carbon varied from 16% to 53% indicating that black carbon constitutes a significant pool of sedimentary organic carbon in the Thracian sea. A fraction ranging from 18% to 97% was scored as nucleoid containing cells. We did not record any significant differences in the fraction of nucleoid-containing bacteria among sediment depths (P<0.05) indicating that there was no accumulation of dead bacterial cells with depth. The same was observed for the fraction of black carbon and bioavailable organic carbon with sediment depth (P<0.05) indicating that benthic consumers are not the key regulators of the organic matter pool in these sediments but have a minor effect. A possible reason for these observations and for the uncoupling between the active bacterial fraction and the bioavailability of organic matter could be (i) the presence of refractory components in the estimated bioavailable organic matter and (ii) the hydrological and geological complexity of the study area. The North Aegean marginal slopes are highly unstable experiencing frequent seismic events. These events are capable of inducing sediment transport from the upper slopes thus altering the entire sediment profile. On the other hand, the significant correlations that were recorded between nucleoid-containing cells and phytopigments (chlorophyll a, phaeopigments, chloroplastic pigment equivalents) at all sediment depths indicate that bacterial communities respond immediately to the deposited phytodetritus, using it as a primary source of carbon and energy. Our data suggests that the Thracian Sea sediments are by no means homogeneous and can best be described as a mosaic controlled by numerous local and regional environmental factors.     

The large penumbra: Long-distance effects of artificial beach nourishment on Posidonia oceanica meadows

We used modelling and field approaches to assess the influence of beach nourishment on a relatively distant Posidonia oceanica seagrass meadow in the NW Mediterranean. Both sediment transport models and in situ observations showed that, two years after the nourishment and under wave climates prevalent during the study period, sediment movement was restricted to shallow waters (<7 m), above meadow shallow limit. The only significant impact on seagrass meadows during this period was an increase in fine sediments, associated with vertical rhizome growth rates 1.5–1.7 times higher than normal. However, running the model with data of wave climate over several decades indicates that strong storms able to transfer these sediments much deeper, potentially burying meadows, occur with a return period of about 25 years. Taken together, our results suggest that beach nourishment could result in significant sub-lethal and lethal consequences for seagrasses that may go unnoticed with short-term evaluations.     

Acute toxicity of mine tailings to four marine species

Acute toxicity bioassays were conducted on mine tailings produced by pilot plant testing for the proposed Quartz Hill molybdenum mine, which will be situated near Ketchikan, Alaska. Tailings bioassays were conducted in seawater with juvenile coho salmon (Oncorhynchus kisutch), mussel larvae (Mytilus edulis), infaunal amphipods (Rhepoxynius abronius), and euphausiids (Euphausia pacifica). The same general range of mine tailings concentrations was acutely toxic to all four test species with acute effects observed between 61 000 to 277 000 mg l^?1 (wet wt) tailings solids (range of 95% confidence limits for LC₅₀ and EC₅₀ values). Chemical analyses of bioassay test solutions and leaching test solutions were conducted for metals (including Cd, Cu, Pb, Zn, Mn and Mo), EPA Priority Pollutant base/neutral organics, and more general parameters such as sulphate, nitrate/nitrite, cyanides, phosphate and ammonia. Parameters possibly contributing to the observed toxicity were complex contaminant mixtures including total suspended solids and heavy metals. The present study provides information related to the marine disposal of mine tailings and shows that these mine tailings present a relatively low level of acutely toxic effects.     

Validation of Arenicola marina in field toxicity bioassays using benthic cages: biomarkers as tools for assessing sediment quality

Sediment toxicity assessments using caged organisms present advantages over using laboratory and native community studies. The use of caged Arenicola marina in sediment toxicity assessments was evaluated. Lugworms were exposed in situ to sediments from coastal and port areas in Spain for seven days, and the activities of the biotransformation enzymes ethoxyresorufin O-deethylase, dibenzylfluorescein dealkylase and glutathione S-transferase, the activities of the antioxidant enzymes glutathione reductase and glutathione peroxidase and lipid peroxidation were then analyzed as biomarkers. Biomarker results and sediment physicochemical data were integrated. Cádiz Bay (SW Spain) sediments presented metal contamination that was not linked to a biochemical response. In LPGC Port (SW Spain), Pb contamination exhibited a moderate toxic potential, while PAHs, and presumably pharmaceuticals, provoked biochemical responses that efficiently prevented lipid peroxidation. In Santander Bay (N Spain), exposure to PAHs and, presumably, pharmaceuticals induced biomarker responses, but lipid peroxidation occurred nevertheless. These results indicated that caged A. marina were effective for the assessment of sediment quality and that the selected biomarkers were sufficiently sensitive to identify chemical exposure and toxicity.     

Sublethal effects of heavy metal contaminated sediments on the bivalve Macoma balthica (L.)

Sublethal effects of estuarine sediments containing high levels of heavy metals were studied using the marine bivalve, Macoma balthica. Burrowing behaviour was inhibited in all contaminated sediments compared to the control. The time required for 50% of the population to burrow (ET50) ranged from 0.17 h in the control to 4.8 h in the most contaminated substrate. A comparison of linear regressions of the concentrations of individual metals in the sediments vs the burrowing response times showed the regressions for mercury and cadmium to be significant (P < 0.05 and < 0.001 respectively). An active threshold avoidance response by burrowed M. balthica was also demonstrated as clams showed a significant (P < 0.05) avoidance of the sediment containing the highest metal levels. Both behavioural responses were considered useful sublethal tests to assess the impact of polluted sediments.     

Evaluation of induction of metallothionein-like proteins (MTLPs) in the polychaetes for biomonitoring of heavy metal pollution in marine sediments

Polychaetes are suitable organisms for evaluation of impact of sediment pollution. We evaluated toxicity of cadmium and copper and measured metallothionein-like proteins (MTLPs) in the polychaete Perinereis nuntia. At the same concentration ranges copper was unexpectedly more toxic than cadmium. Copper also caused no significant increase in MTLPs in the polychaetes. When P. nuntia and another polychaete species, Cirratulus cirratus were cultured in the contaminated sediments collected from Lake Sihwa (Korea), a high mortality of 80% was observed on day 6 in P. nuntia in the sediment with the highest metal concentration. However, no mortality was observed up to 35 days in C. cirratus in any sediment. MTLP contents between two species also varied. These findings suggest that MTLP induction response in the polychaetes varies with the metal type and species and it may be used as a biomarker of sediment pollution in the polychaetes after further validation and field trials.     

Metabolism of a burrowing polychaete: Precaution needed when measuring toxic effects

The technique of direct calorimetry has been used to monitor the metabolic activity of the polychaete, Neanthes virens, an organism commonly used for testing the toxic effects of chemicals in the marine environment. In these tests Neanthes showed unpredictable alternating periods of hyperactivity and rest when deprived of sediment. If used in this abnormal state for toxicity tests, the results would represent the effects of the chemicals on abnormal organisms and may not, therefore, be realistic information for judging the chemical's harmful effects on it and other species in nature. In toxicity tests with heavy metals that have been reported in the scientific literature, the shorter survival time of test organisms without sediment, as compared to those in sediment, was probably the result of the abnormally high metabolic rates of the former group.     

Burrowing responses of the short-neck clam Ruditapes philippinarum to sediment contaminants

The burrowing responses of a common tropical bivalve, the short-neck clam Ruditapes philippinarum, to cadmium (Cd)-spiked sediment, variations of sediment grain size and natural sediments collected from 15 locations in Hong Kong's inshore waters were investigated through a series of laboratory tests. Results showed that the burrowing response exhibited a negative relationship with an increase in Cd concentration in the spiked sediments. The level of Cd was also found to be directly proportional to the percent mortality of the clam. However, there was no significant difference (p>0.05) in the time elapsed for the clam to burrow into sediments with different grain size composition. The elapsed time for 50% of the test clams to burrow into the sediment (ET₅₀) over a period of 48 h was calculated for the sediment samples collected from the 15 locations. Results of ANOVA showed significant difference (p<0.05) among the sediment samples. Tukey's multiple comparison test revealed two groups of sediments: group 1 containing 3 sediment samples collected from Victoria Harbour and group 2 containing 12 samples obtained from other coastal areas of Hong Kong. The ET₅₀ value for group 1 sediments was found to be greater than 2880 min whilst the ET₅₀ for group 2 sediments had a mean of 173.9 min. Agglomerative classification of the sediment samples, based on metal content (As, Cd, Cr, Cu, Ni, Pb, Zn), also showed two similar groups, suggesting that the ET₅₀ values were correlated with the metal level of the sediment samples. Group 1 sediments collected from Victoria Harbour had greater concentrations of Cd, Cr, Cu, Ni and Zn as compared to that in group 2. The present study demonstrated that high metal content in the sediment tends to inhibit the burrowing responses of the clam and that there is potential to develop the burrowing responses of R. philippinarum as a sublethal sediment toxicity test.