Differentiation between bioavailable and total hazard potential of sediment-induced DNA fragmentation as measured by the comet assay with Zebrafish embryos

Goals, Scope and Background  While water quality strongly improved over decades in the Rhine River, sediments still reflect elapsed contaminations of organic pollutants and heavy metals. In comparing genotoxic effects induced by both sediment extracts and whole sediments, a ratio of bioavailable toxicity and total extractable toxicity is obtained. Since contaminated sites whose contaminants are toxic and as well bioavailable present an elevated risk to the ecosystem, such ratios may be used as a warning signal to identify sites of primary concern. Methods  Accordingly, two different exposure scenarios were compared to reveal the genotoxic potential of 18 sediment samples derived from 9 sample sites along the River Rhine. For assessment of effects on genome integrity, DNA fragmentation was measured using the comet assay with primary cells isolated from zebrafish embryos previously exposed to either organic sediment extracts or freeze-dried sediments at sublethal concentrations. Additionally, chemical data were used to determine responsible pollutants and correlate them with biological effects. Results  Whereas 17 out of 18 sediment extracts caused significant DNA damage to the embryo cells, only 4 native sediments showed a genotoxic potential. Thus, under field-like exposure conditions, a major part of potentially genotoxic compounds seem to remain particle-bound and ineffective, as shown for whole sediment exposure. Conversely, the organic extracts seem to contain enriched concentrations even of hardly soluble substances. Hence, organic extracts may be used as a screening tool to address potentially polluted sites, even though the relevance of these results for the field situation may be questionable. Investigations on native sediments determined few sites with bioavailable and therefore ecologically most relevant genotoxic sediment compounds. Discussion  However, these results may underestimate the total hazard potential of sample sites with hardly bioavailable substances. Chemical data revealed a variety of anthropogenic pollutants, ranging from PAHs to heavy metals. Nevertheless, chemical data on the measured priority pollutants did not fully explain the pollution pattern of the bioassays but clearly determined substances of concern (e.g., HCB, heavy metals) in particular sample sites. Conclusions  There is a striking advantage in assessing the genotoxicity by means of different exposure scenarios that focus on either bioavailable or extractable fractions, as the combination of the results allows obtaining information on specific properties of the genotoxicants and their bioavailability. An additional correlation with chemical data should be required to identify priority pollutants, as long as the responsible contaminant is known a priori. As many studies revealed inherent failures of such a correlation, an effect-driven analysis of pollutants is recommended as a promising tool to identify even non-priority pollutants by means of their ecotoxicological effectiveness.     

Hazard identification of contaminated sites—ranking potential toxicity of organic sediment extracts in crustacean and fish

Background, aim, and scope  It is well known that contaminated sediments represent a potential long-term source of pollutants to the aquatic environment. To protect human and ecosystem health, it is becoming common to remediate contaminated sites. However, the great cost associated with, e.g., dredging in combination with the large numbers of contaminated sites makes it crucial to pinpoint those sites that are in greatest need of remediation. In most European countries, this prioritization process has almost exclusively been based on chemical analyses of known substances; only seldom toxicity data has been considered. The main objective of the current study was therefore to develop a tool for hazard identification of sediment by ranking potential toxicity of organic sediment extracts in a crustacean and a fish. A secondary objective was to investigate the difference in potential toxicity between compounds with different polarities. Materials and methods  Early life stages of the crustacean Nitocra spinipes and the fish Oncorhynchus mykiss, which represent organisms from different trophic levels (primary and secondary consumer) and with different routes of exposure (i.e., ingestion through food, diffusive uptake, and maternal transfer), were exposed to hexane and acetone fractions (semi-polar compounds) of sediment from five locations, ranging from heavily to low contaminated. Preliminary tests showed that the extracts were non-bioavailable to the crustacean when exposed via water, and the extracts were therefore loaded on silica gel. Rainbow trout embryos were exposed using nano-injection technique. Results and discussion  Clear concentration–response relationships of both mortality and larval development were observed in all tests with N. spinipes. Also for rainbow trout, the observed effects (e.g., abnormality, hemorrhage, asymmetric yolk sac) followed a dose-related pattern. Interestingly, our results indicate that some of the locations contained toxic semi-polar compounds, which are normally not considered in risk assessment of sediment since they are focused on compounds isolated in the hexane fraction. Conclusions  The ranking of the five sediments followed the expected pattern of potential toxicity in both organisms, i.e., sediments with known pollution history caused major effects while reference sediments caused minor effects in the two test systems. Silica gel turned out to be an excellent carrier for exposure of N. spinipes to very hydrophobic and otherwise non-bioavailable sediment extracts. Recommendations and perspectives  Since both test systems demonstrated that a substantial part of the potential toxicity was caused by semi-polar compounds in the acetone fractions, this study enlightens our poor understanding of which compounds are causing adverse effects in environmental samples. Therefore, by investigating potential toxicity (i.e., hazard identification) as a first screening step in prioritizing processes, these implications could be avoided. For proper sediment risk assessment, we however recommend whole sediment toxicity tests to be used for selected sites at following tiers.     

Polycyclic aromatic compounds and microtox® acute toxicity in contaminated sediments in Sweden

Limnic and brackish aquatic sediments from contaminated locations in Sweden were analysed for polycyclic aromatic compounds (PACs) and tested for acute aquatic toxicity. The organic compounds were analysed in solvent-extractable and alkaline-treated fractions to complete the analysis of a set of priority pollutants according to the Swedish EPA. Additionally, the acute toxicity was measured by the solid phase Microtox test. The measured effects were correlated with sedimentological and chemical parameters. Analysis revealed no indication of a contribution of PACs to the acute sediment toxicity in highly contaminated sediments, with concentrations in the range of 11.3-307 μg SPAC_37/39/g dry matter. Despite a high ranking of 3 to 5 within the Swedish EPA list for coastal sediments, the acute toxicity results indicate a very low bioavailability of the analysed PACs from the creosote and combustion-contaminated sediments. A correlation of acute toxicity to elemental sulfur was indicated.     

Contaminated sediments as a potential source of Zn, Pb, and Cd for a river system in the historical metalliferous ore mining and smelting industry area of South Poland

Background, aim, and scope  Elevated levels of heavy metals in the aquatic and soil systems can be caused by the weathering of mineralized rocks. This enrichment is often considerably enlarged by historical and current mining and smelting activities. In Poland, the most contaminated river systems are those in the Silesia region. The metalliferous ore mining and smelting industries have been the main sources of heavy metal pollutions over the last 100–170 years. The previous and present studies have shown very high concentrations of heavy metals in the bottom sediments of the Mala Panew River, the most polluted tributary of the Oder River. The main objective of this work was to study temporary changes of selected metal (Zn, Pb, and Cd) concentrations in upper layer of bottom sediments at the measuring point near the outlet of the Mala Panew River into the Oder River, and to determine the vertical distribution of the metals in the sediment cores from the most polluted middle part of this river. The mobility of the metals and their potential bioavailability were assessed based on metal partitioning in the sediments and metal concentrations in pore waters. The presented data were compared with metal concentrations in aquatic sediments from similar historical mining and smelting sites in Poland and other countries. Methods  The upper layer of bottom sediment samples from the same Mala Panew River measuring point were collected six times in the period 1997–2005, while five sediment cores were collected once from the middle course of Mala Panew River in 2006. Abiotic parameters such as pH and Eh have been determined in situ. Metal contents were determined in the <20 and <63 μm size fractions of sediments after digestion in a microwave oven with aqua regia or concentrated nitric acid. Metal mobility was assessed in the selected sediment cores by the chemical forms of metals (sequential extraction method) and their concentrations in pore waters were investigated. Results  The concentrations of Cd, Pb, and Zn in the upper layer of sediments varied, depending on both the season and the year of sampling. Their mean concentrations (from six samplings) are [mg/kg]: Zn 1,846, Pb 229 and Cd 73. The metal concentrations in the sediment cores varied with the depth in the range of [mg/kg]: 0.18–559 for Cd, 26.2–3,309 for Pb and 126–11,153 for Zn, although the highest accumulations generally could be observed in the deeper layers. The most mobile metal fractions, i.e., exchangeable, carbonate and easily reducible fractions, are typical of Zn and Cd. Cadmium was found to be the most mobile metal and its relative contribution ranges from 84 to 96%, while in the case of Zn it ranged from 45 to 94%. Lead is mainly associated with the moderately reducible fraction (30–60%). Relative contributions of metal chemical forms slightly vary with the depth in the sediment profile. The results obtained for the pore water samples show very high concentrations of the metals studied, especially in the case of Cd (31–960 μg/dm³) and Zn (300–4,400 μg/dm³). Discussion  Accumulation of Cd, Pb, and Zn in the upper layer of the bottom sediments and in the sediment core samples from the Mala Panew River is very high, considerably exceeding the local geochemical background. High contributions of mobile Cd and Zn and the toxicity of cadmium can cause environmental risk. Our measurements also suggest that mobile metals can migrate into groundwater, whereas the groundwater itself can leach some chemicals from river sediments, because of a relatively high water table in the study area, especially during rainfall periods. Comparison of the results obtained with the literature data from the last decade shows that the concentrations of Cd and Zn in the sediments from the Mala Panew River are the highest among other submersed sediments in Poland and other regions (e.g., the Mulde River, Germany). Conclusions  The Mala Panew River is one of the most polluted rivers when compared with similar historical mining and smelting areas in Poland and elsewhere. The sediments studied are strongly polluted with the metals analyzed. In the upper layer of the bottom sediments there has been no reduction of Zn and Cd amounts over the last decade, which could suggests a long-term migration and a secondary contamination. Considerably higher accumulations of metals in overbank sediment cores and in the deeper core section could result from strong contamination in previous decades and translocation of Cd and Zn (secondary pollutants). The relatively high concentrations of the two metals in pore waters support these findings. Cadmium is crucial in the environmental risk assessment because of its high mobility and toxicity. These data are important for water/sediment management in the transboundary Oder River catchment, situated in Poland, Germany and the Czech Republic. Recommendations and perspectives  It is important to assess mobility phase and pore water in the contaminated historical aquatic sediments. Such studies may help explain the changes, which take place in the sediment layers as well as at the water–sediment interface. Obtained results should be used for the risk assessment of the historical contaminated sediments at the local river-basin scale. The treatment of contaminated sediments, e.g., dragging activity, should be considered as very important in management strategies in order to avoid remobilization of metals.     

Importance of dose metrics for lethal and sublethal sediment metal toxicity in the oligochaete worm Lumbriculus variegatus

Background, Aims, and Scope  There is an increasing demand for controlled toxicity tests to predict biological effects related to sediment metal contamination. In this context, questions of metal-specific factors, sensitivity of toxicity endpoints, and variability in exposure duration arise. In addition, the choice of the dose metrics for responses is equally important and is related to the applicability of the concept of critical body residue (CBR) in exposure assessments, as well as being the main focus of this study. Methods  Experiments were conducted to assess toxicity of Cd, Cr, Cu and Pb to the oligochaete worm Lumbriculus variegatus with the aim of determining CBRs for two response metrics. Mortality and feeding activity of worms exposed to sediment-spiked metals were used as end-points in connection with residue analyses from both the organisms and the surrounding media. Results  LC50 values were 0.3, 1.4, 5.2, and 6.7 mg/L (from 4.7 μmol/L to 128.0 μmol/L), and the order of toxicity, from most toxic to least toxic, was Cu > Cd > Pb>Cr. By relating toxicity to body residue, variability in toxicity among the metals decreased and the order of toxicity was altered. The highest lethal residue value was obtained for Cu (10.8 mmol/kg) and the lowest was obtained for Cd (2.3 mmol/kg). In the 10-d sublethal test, both time and metal exposure were an important source of variation in the feeding activity of worms. The significant treatment effects were observed from worms exposed to Cd or Pb, with the controls yielding the highest feeding rate. However, quantitative changes in the measured endpoint did not correlate with the exposure concentrations or body residues, which remained an order of magnitude lower than in the acute exposures. Discussion  Both response metrics were able to detect a toxic effect of the metals. However, the ranking of metal toxicity was dependant on the choice of the dose metric used. An attempt to form a causal mortality-mediated link between tissue residues and metal toxicity was successful in water-only exposures. The results also indicated that egestion rate was a sensitive toxicity end point for predicting the effects of sediment contamination. Conclusions  By relating the biological response with the tissue metal residues, toxicity data was comparable to both environmental media as well as different response metrics and time scales. The results also revealed the importance of metal toxicity ranking on a molar basis and, furthermore, a direct link to the CBR concept was established. Recommendations and Perspectives  There is a growing demand for methods to assess the effects of contaminated sediments to benthic fauna and whole aquatic ecosystems. Such information is needed for sediment quality guidelines that are currently being developed in many countries and remediation processes. The use of body residues as a dose metric in metal toxicity studies may help to overcome difficulties related to bioavailability issues commonly faced in sediment toxicity studies. ESS-Submission Editor: Prof. Dr. Henner Hollert (henner.hollert@bio5.rwth-aachen.de)     

Characterization and Degradation of Petroleum Hydrocarbons Following an Oil Spill into a Coastal Environment of South Texas, U.S.A.

Petroleum hydrocarbons were characterized at eleven sites withinthe sediments of a coastal stream in south Texas, U.S.A. following a medium sized crude oil spill. Bank and open-water(deep) sediments were collected at each site. Hydrocarbonstargeted for analysis included 22 aliphatic hydrocarbons (C-11to C-34) and 16 polynuclear aromatic hydrocarbons. Sedimentconcentrations were measured at intervals of 1, 2, 3, 4, 6, and12 months post-spill. Higher hydrocarbon concentrations were observed for a longerduration within the deep sediments than bank sediments. Initialhydrocarbon constituents in impacted sediments matched the crudeoil fingerprint accurately with the exception of the lighter-endhydrocarbons. The lighter-end aliphatic hydrocarbons areaffected immediately by evaporation and dissolution processesduring the spill event and were found below the detection levelsat most of the sites. Total hydrocarbon concentrations insediments within each hydrocarbon group returned to backgroundlevels by the end of the study period. Observed decreases inhigh molecular weight polycyclic aromatic hydrocarbons (PAH)concentrations exceeded known environmental degradation rateswhich suggests the influence of a sediment transport process. Overall, the fate of petroleum hydrocarbons within this type ofenvironment were likely related to both degradation and sedimenttransport processes. By the end of the study period, most individual PAH constituent concentrations were below thresholdconcentrations thought to produce toxic effects in marine andestuarine organisms. PAH constituents concentrations remainingabove threshold concentrations included benz(a)anthracene,chrysene, and benzo(a)pyrene.     

A Novel Particle Contact Assay with the Yeast Saccharomyces cerevisiae (8 pp)

Goal, Scope and Background   Numerous xenobiotics released into surface waters are transferred to suspended particulate matter and finally attached to sediments. Aquatic organisms may be exposed to them by direct particle feeding, by physical contact with contaminated surfaces as an exposure route, and by the uptake of dissolved contaminants after equilibration via the free water phase. In order to assess potential sediment toxicity, each of these exposure routes has to be addressed. This paper presents a newly developed particle contact assay that uses the fermentation performance of a specific Saccharomyces cerevisiae strain for the assessment of toxic effects in sediments. The test procedure is based on the characteristic feature of growing yeast cells to attach to sediment particles, which are also relevant for the accumulation of contaminants. The physical contact with lipophilic contaminants mirrors an exposition pathway for the direct uptake into the cells. In order to quantitatively characterize the toxic effects of particle attached pollutants on the fermentation performance, unpolluted native reference sediment was spiked with representatives for widely distributed anthropogenic contaminants. Methods   Saccharomyces cerevisiae was established as sensitive eukaryotic microorganism for the ecotoxicological assessment of particle attached anthropogenic contaminants in freshwater sediments. For this purpose, yeast cells were cultivated in sediment samples and the resulting fermentation performance was continuously measured. Sediments artifically spiked with HCB, PCB, g-HCH, DDT, and benzo(a)pyrene and solutions of each contaminant were comparatively investigated by means of their adverse effects on yeast fermentation performance. Additionally, four native river sediments characterized by increasing levels of pollution were assessed by the yeast particle contact assay, and simultaneously by standard aquatic tests with algae, daphniae, and luminescent bacteria using pore water and elutriates. Results of the bioassays were related to specific sediment contamination with respect to metals and organic priority pollutants. Results and Discussion   In sediments spiked with PCB and benzo(a)pyrene fermentation, performance was affected extensively below concentrations inhibiting fermentation in contaminant solutions. This suggests a high efficiency of the exposure route by physical contact. The fermentation performance was only slightly affected by single lipophilic pollutants, whereas mixtures of individually spiked sediments caused critically reduced fermentation performance suggesting additive synergistic effects. Native river sediments modestly to critically polluted by hazardous organic compounds lead to a slightly to dangerously reduced fermentation performance in the yeast contact assay. These inhibitory effects were much less pronounced in the standard bioassays conducted with algae, daphniae and luminescent bacteria, applying pore waters and elutriates as sample matrices. Using pore water, inhibition was measured only in the most polluted sediment, elutriates lead to a slight inhibition of the algal growth in the undiluted sample only. These results indicate an improved sensitivity of the yeast particle contact assay compared to the standard assays, due to uptake and physical cell contact as additional routes of exposure. Conclusion   The yeast particle contact assay is a valuable tool for the assessment of ecotoxicological potential in freshwater sediments. Since the assay addresses physical contact as an exposure route, it indicates bioavailability of lipophilic compounds in sediments. Outlook   The sensitive indication of bioavailable contaminants associated to sediment particles by the newly developed yeast particle contact assay recommends it as a complementary microbial bioassay in a test battery for assessing major pathways of contaminants in whole sediments.     

Rapid sedimentological and biological assessment of hydrocarbon contaminated sediments

It is possible to rapidly detect the presence of high concentrations of sediment associated hydrocarbons using a sediment profile camera and simultaneously evaluate the general sedimentological and biological character of a contaminated area. In sediments that were heavily contaminated with hydrocarbons from spills and chronic long-term additions the presence of hydrocarbons was seen about 50% of the time in the sediment profile images as unique features, ‘H spots’. The presence of these features was related to the concentration of hydrocarbons in the sediment. In highly contaminated muddy sediments ‘H spots’ were found in images collected at stations that had from 270 to 610 ppt total hydrocarbons. Sedimentological and biological information obtained from the sediment profile images confirmed the impacted nature of Elizabeth River sediments. Sediment profile imaging provide a means of obtaining an overall evaluation of the quality of a habitat and impacts on that habitat from pollution related environmental disturbances. While qualitative, an advantage of sediment profile image data is that they can be evaluated in less than a day and used to quickly locate inclusions of hydrocarbons in the sediments for further quantitative chemical or biological sampling, or mapping of heavily contaminated areas.     

Ecotoxicological risk assessment of a quarry filling with seaport sediments using laboratory freshwater aquatic microcosms

Purpose

In France, contaminated seaport sediments cannot be discharged into the sea according to recent regulation. Hence, they must be managed on land. Among the solutions identified, that of filling terrestrial quarries located in the littoral zone is one of the most promising. However, this requires developing a methodology for ecological risk assessment; which was the aim of the SEDIGEST research program. In the present study, we report the results of toxicological impacts of three sediments on aquatic ecosystems close to a quarry. These results were used to formulate a risk assessment methodology.

Materials and methods

The ecotoxicological approach was based on laboratory microcosm assays applied to leachates obtained from the sediments. The microcosms contained synthetic water and sediment and five pelagic (micro-algae, duckweeds and daphnids) and benthic (chironomids and amphipods) freshwater species. The biota were exposed for 3 weeks to a range of leachate concentrations; metals were monitored in the water column and the effects (i.e. mortality, growth inhibition and midge emergence) were measured.

Results and discussion

The results showed an absence of acute toxicity at concentrations of <10% (v/v) but sublethal effects for concentrations between 1 and 10%. Given the complex nature of the leachates, it was difficult to determine the factors of toxicity. Metals, especially Cu, might have been responsible for some of the effects on the amphipods.

Conclusions

Future quarries filled with seaport sediments might emit pollutants to aquatic ecosystems in their vicinity, and risk assessors should ensure that predicted environmental concentrations of leachates are below the maximum leachate concentration acceptable for the whole ecosystem; for example, by multiplying the concentration safe for the microcosm organisms by a factor of 10.     

Contribution of Oil Industry Activities to Environmental Loads of Heavy Metals in the Tabasco Lowlands, Mexico

In Tabasco the petroleum industry pollutes soil recurrently by oil spills. We analysed Pb, V, Ni and Cr concentrations in water samples, and total metal contents and metal fractions in soil samples of contaminated and non-contaminated soils and in sediments. Besides, we determined Eh, pH, DOC and major ions in water and Eh, pH, C_org in soils and sediments. Sediments contained considerably larger heavy metal (HM) concentrations than soils. Local background concentrations of V, Ni and Cr in soils are larger than global means and oil spillages have not added these metals in quantities that exceed the natural variation. Spillage of formation water increases Pb concentrations in soils, particularly in mobile fractions. The contribution of the oil industry to HM loads is diluted by large fluvial water and sediment discharges and difficult to assess by comparison of total metal contents. Therefore, easily mobile metal fractions are much better indicators.     

A new sediment contact assay to assess particle-bound pollutants using zebrafish (<Emphasis Type="Italic">danio rerio</Emphasis>) embryos

Goal, Scope and Background. Based on a bioassay battery covering only primary producers and consumers as well as degraders, the potential ecological hazard of sediments to vertebrates cannot be estimated comprehensively. Therefore, there is an urgent need to develop and standardize integrated vertebrate-based test systems for sediment investigation strategies. Whereas vertebratebased in vitro systems have frequently been used for the investigation of aqueous samples, there is a significant lack of whole sediment assays. Thus, the purpose of the present study was: (1) to develop a rapid and reliable, but comprehensive method to investigate native sediments and particulate matters without preceding extraction procedures; (2) to compare the hazard potential of solid phase sediments to the effects of corresponding pore waters and organic extracts in order to characterize the bioavailability of the particle-bound pollutants; and (3) to relatively evaluate the embryotoxic effects of sediments from the catchment areas of the rivers Rhine, Neckar and Danube. Methods (or Main Features).  To investigate the toxicity of sediment samples on vertebrates, the standard embryo toxicity test with the zebrafish (Danio rerio; Hamilton-Buchanan 1922) according to DIN 38415-6 was modified with respect to exposure scheme and toxicological endpoints. Sediments from the catchment area of the Neckar River were assessed using pore waters, acetonic extracts and native sediments in order to get inside into the potential bioavailability of particle-bound pollutants. A comprehensive test protocol for the investigation of native sediments in the embryo toxicity test with the zebrafish is presented. Results and Discussion.  The fish embryo assay with Danio rerio can be carried out with both aqueous and organic sediment extracts as well as native (whole, solid phase) sediment samples. Elongation of exposure time from 48 to up to 196 h significantly increased the mortality. Using the fish egg assay with native sediments, a broad range of embryotoxic effects could be elucidated, including clear-cut dose-response curves for the embryotoxic effects of contaminated sediments; in contrast, absence of embryotoxic effects could be demonstrated even for the highest test concentrations of unpolluted sediments. With native sediments, embryotoxicity was clearly higher than with corresponding pore waters, thus corroborating the view that — at least for fish eggs — the bioavailability of particle-bound lipophilic substances in native sediments is higher than generally assumed. The relative ranking of sediment toxicity was identical using both native sediments and sediment extracts, EC₂₀ values of the latter, however, being eight time lower higher than with the native sediments. A comparison of the embryo toxic effects of samples from the Neckar area with locations along the Rhine and Danube rivers elucidated a broad range of results, thus indicating different levels of contamination. Conclusions.  A modified protocol of the zebrafish embryo test allows the assessment of sediment toxicity in both aqueous extracts and native sediments. The isolated investigation of pore waters may result in a clear-cut underestimation of the bioavailability of lipophilic particle-bound substances (as determined by native sediments). Recommendations and Perspectives.  The zebrafish embryo test with native (whole, solid phase) sediments appears very promising for the evaluation of the bioavailable fraction of lipophilic particle-bound substances and can therefore be recommended for the evaluation of vertebrate toxicity in tiered sediment test strategies and dredging directives such as the HABAB-WSV. Whereas acetone extracts may be tested as a rough estimation of embryotoxicity, native sediment samples will provide a more comprehensive and realistic insight into the bioavailable hazard potential     

Evaluation of a Multi-species Test System for Assessing Acute and Chronic Toxicity of Sediments and Water to Aquatic Invertebrates: Effects of Pentachlorophenol on Daphnia magna and Chironomus prasinus (6 pp)

Background, Aims and Scope   Single-species toxicity tests are currently used as main tools for assessing effects in ecological risk assessments. Nevertheless, in higher-tier evaluations, alternative ecotoxicological methods can be required. Alternative methods should cover more realistic exposure conditions and/or additional species or endpoints. Water/sediment systems can be considered for covering additional exposure routes. A typical feature of water/sediment systems is the modification of exposure levels with time. The simultaneous exposure of different species within the same system can cover differences in sensitivity and relevance of exposure routes. The aim of this study was to develop a water/sediment test that could detect relevant effects on two selected species and explore the relevance of sediment and waterborne exposure routes. This alternative method may provide relevant information facilitating the design of microcosm and mesocosm studies.Methods   A multi-species water/sediment system was designed using two invertebrates: Chironomus prasinus a benthic detritivore invertebrate and Daphnia magna a pelagic filtering invertebrate was designed. Seven static water/sediments systems were built up: two controls and five treatments where sediments were spiked with NaPCP at nominal concentrations of 1.25, 2.5, 5, 10 and 20 mg/kg, respectively. Each system contained caged and free (unrestrained) Daphnia magna organisms to consider differences in the exposure route (through sediment and/or water column) and Chironomus prasinus organisms. Fate and partitioning of PCP within the water/sediment compartments were assessed. Acute (lethality) and chronic effects (reproduction) for Daphnia magna were monitored after 48 h and 16 days of exposure, respectively. Adult emergence and oviposition success of Chironomus prasinus were monitored at the end of test (16 days).Results and Discussion   Mortality and reproduction inhibition of Daphnia magna occurred at the two higher doses, corresponding to maximum measured PCP water concentrations of 1.95 and 0.746 mg/l, respectively. Chironomus prasinus was less sensitive to PCP than Daphnia magna. Concentrations inhibiting reproduction and provoking almost 100% adult mortality in D. magna only reduced slightly the emergence and reproduction of chironomids. The experimental design focussed on suitability of the system for assessing simultaneously assessing effects on D. magna and C. prasinus instead of calculations of toxicity endpoints for PCP. The results indicate that this experimental design can be useful to the field of sediment/aquatic toxicity testing, particularly through the comparison with single- species results. Assessment of relative contribution of two exposure routes for Daphnia magna and complementary chronic endpoints derived from two different species can be achieved. Another advantage is that differences in reproduction strategies of selected species (sexual vs. parthenogenetic) should permit one to detect mechanisms of action associated to with endocrine disrupters.Conclusion   The feasibility of a two-species test built-up in an artificial sediment/water system has been demonstrated. This experimental design represents a cost/effective test which can assess effects through water and sediment pathways and incorporates several relevant chronic endpoints in a single assay. Recommendation and Outlook   By building on the experience gained, improvements for further test designs are discussed. Test validation with a set of model chemicals will contribute to address the effectiveness of the test developed as comparing compared to single- species tests.     

Analysis of burrowing ability as a sublethal endpoint in a marine sediment bioassay with Corophium volutator (Pallas)

Purpose

Acute whole-sediment bioassay with the estuarine and marine amphipod Corophium volutator (Pallas) is widely used to assess toxicity of sediments. According to the guidelines DIN EN ISO 16712, mortality is the determined toxic endpoint. Additionally, the reburrowing ability of the surviving organisms of this acute toxicity test in fresh uncontaminated sediment is suggested as the sublethal endpoint, but insufficient information (e.g., exact measurement protocols) on this endpoint is provided, thus confounding factors and the interpretation of the results. The aim of this study was to provide information on burrowing activity as a sublethal endpoint.

Materials and methods

Amphipod tests were carried out in the laboratory, and the burrowing behavior was examined in a size- and gender-specific manner. For sediment testing, only animals of the same size were used in a defined sex ratio because it was found that female animals buried themselves faster than males and that smaller animals burrowed faster than bigger organisms. Statistical analyses were applied to determine whether burrowing time and ability differ significantly between sexes and sizes. Finally, tests were run to discern whether the burrowing ability could be a more sensitive endpoint than mortality.

Results and discussion

When the burrowing ability was examined in toxicity tests with contaminated sediment, the test organisms were affected in a dose-dependent manner. With rising concentrations of the contaminated sediment in a sublethal testing following the sediment exposure over 10 days, fewer animals buried themselves into the sediment.

Conclusions

The burrowing behavior can be used as an additional endpoint. For the tested sediment, burrowing was found to be more sensitive than the mortality. Guidance on the measurement protocol for this additional endpoint was developed. Under the test conditions examined, burrowing ability is an appropriate sublethal endpoint to supplement the toxicity test procedure.     

Field-Scale Assessment of Phytotreatment of Soil Contaminated with Weathered Hydrocarbons and Heavy Metals (9 pp)

Background, Aim and Scope   Phytoremediation is a remediation method which uses plants to remove, contain or detoxify environmental contaminants. Phytoremediation has successfully been applied for the removal of fresh hydrocarbon contamination, but removal of aged hydrocarbons has proven more difficult. Biodegradation of hydrocarbons in the subsurface can be enhanced by the presence of plant roots, i.e. the rhizosphere effect. Phytostabilization reduces heavy metal availability via immobilization in the rhizosphere. Soils contaminated by both hydrocarbons and heavy metals are abundant and may be difficult to treat. Heavy metal toxicity can inhibit the activity of hydrocarbon-degrading microorganisms and decrease the metabolic diversity of soil bacteria. In this experiment, weathered hydrocarbon- and heavy metal- contaminated soil was treated using phytoremediation in a 39- month field study in attempts to achieve both hydrocarbon removal and heavy metal stabilization. Materials and Methods: A combination of hydrocarbon degradation and heavy metal stabilization was evaluated in a field-scale phytoremediation study of weathered contaminants. Soil had been contaminated over several years with hydrocarbons (11400±4300 mg kg dry soil)-1 and heavy metals from bus maintenance activities and was geologically characterized as till. Concentrations of soil copper, lead and zinc were 170±50 mgkg-1, 1100±1500 mg kg-1 and 390±340 mg kg-1, respectively. The effect of contaminants, plant species and soil amendment (NPK fertilizer or biowaste compost) on metabolic activity of soil microbiota was determined. Phytostabilization performance was investigated by analyses of metal concentrations in plants, soil and site leachate as well as acute toxicity to Vibrio fischeri and Enchtraeus albidus. Results: Over 39 months hydrocarbon concentrations did not decrease significantly (P=0.05) in non-amended soil, although 30% of initial hydrocarbon concentrations were removed by the last four months of study. In soil amended with NPK fertilizer and municipal biowaste compost, 65 % and 60 % of hydrocarbons were removed, respectively. The soil contained metabolically diverse bacteria, measured as carbon source utilization and extracellular enzymatic activities. Compost addition resulted in a slight increase in enzymatic activities. Diesel fuel utilization potential in Biolog MT2 plates inoculated with a soil suspension was enhanced by both compost and NPK compared to non-amended soil. Soil toxicity to V. fischeri and E. albidus was low. The leachate was not toxic to V. fischeri. Pine (Pinus sylvestris), poplar (Populus deltoides x Wettsteinii), grasses and clover (Trifolium repens) survived to varying degrees in the contaminated soil. All plants suffered from phytotoxicity symptoms and some trees died during the study period. Plants formed a dense cover over the compost-amended soil, whereas non-amended soil had areas devoid of vegetation throughout the study. Vegetation coverage in the NPK-amended quarter was about 50 % after the first four months of study, but increased gradually to 100 %. Heavy metals did not accumulate in plant tissue. Discussion: Removal of hydrocarbons from weathered unfertilized hydrocarbon-contaminated soil was not statistically significant despite the presence of a viable hydrocarbon-degrading microbial community. This effect is attributed to soil heterogeneity and low bioavailability of hydrocarbons. Hydrocarbon concentrations were not reduced to the desired level, i.e., 1500 mg hydrocarbons (kg of dry soil)-1, in any treatment. . The presence of clay minerals and organic matter within the compost may have limited heavy metal transfer to leachate and plant tissue. Conclusions: Weathered hydrocarbons were partly decomposed in soil fertilized with NPK fertilizer or biowaste compost, but not from unfertilized soil. The active hydrocarbon-degrading microbiota and low toxicity of soil to V. fischeri and E. albidus indicates low availability of contaminants to microorganisms. Despite high heavy metal concentrations, the soil contained metabolically diverse bacteria, measured as carbon source utilization and extracellular enzymatic activities. Heavy metals did not accumulate in test plants. Pine and poplar suffered from phytotoxicity symptoms in the soil and could not enhance hydrocarbon removal in compost-amended soil. Compost addition combined with a grass and legume crop is suggested for stabilization of combined hydrocarbon- and metal-contaminated soil. Recommendations and Perspectives: Both compost and NPK fertilizers can be used to enhance phytoremediation of soil contaminated with weathered hydrocarbons in the presence of heavy metals; however, compost addition is recommended since it enables greater vegetative coverage. This in turn may decrease heavy metal mobility. Phytoremediation can be used for remediation of soil contaminated with weathered hydrocarbons in the presence of heavy metals. However, phytoremediation of weathered contaminants requires extended periods of time; thus, other remediation methods should be considered in the event of soil contamination posing an immediate public health and/or environmental threat.     

The use of <Emphasis Type="Italic">Daphnia magna</Emphasis> immobilization tests and soil microcosms to evaluate the toxicity of dredged sediments

Purpose  

This paper evaluates the feasibility of using the buffering capacity of natural soil for the remediation of dredged material before being disposed in soil landfills. To achieve that, an Integrated Soil Microcosms (ISM) system was designed to produce elutriates and leachates from the sediment/soil percentage mixtures. Furthermore, to investigate the biological effects of the contaminated sediments, the toxicity behavior of leachates and elutriates was assessed and compared by performing acute (48 h) toxicity assays with the cladoceran Daphnia magna as test organism.     

A new plant-based bioassay for aquatic sediments

Background, Goal and Scope  To date, standardised bioassays for the assessment of the ecotoxicological potential in sediments and dredged material use test organisms like bacteria, algae and crustaceae. This paper presents the development and application of a novel sediment contact test (whole sediment) withMyriophyllum aquaticum, a representative of rooted aquatic macrophytes. The aim of the present study is to demonstrate the value of a sediment contact test with rooted macrophytes as a supplement to existing test batteries in order to improve the assessment of sediment toxicity. Methods  The newly developed sediment contact test withMyriophylhim aquaticum was applied to natural whole sediments. For performing the test, whorls ofMyriophyllum aquaticum were directly planted in the native sediment and incubated in the light at 24°C (cf. section results and discussion). The end points of the test were the number of the shoots and the fresh weight of the whole plants. The duckweed growth inhibition test withLemna minor according to ISO/DIS 20079 was performed in pore waters from sediment samples. The results of the sediment contact test withMyriophyllum aquaticum were compared with each other and with those of the aquatic duckweed test. Results and Discussion  A test protocol for the new plant-based sediment contact test using the aquatic plantMyriophyllum aquaticum as an indicator was developed. The best control sediment proved to be the OECD sediment (OECD 207). A test period of 10 days appeared to be sufficient for the test. The increase of biomass and the derived growth rate were found to be the most suitable evaluation parameters. The growth behaviour ofMyriophyllum aquaticum differed depending on the origin of sediments. Therefore, plant-affecting contamination, that is bound in sediments, was indicated. Conclusions  The novel sediment contact test withMyriophyllum aquaticum can indicate phytotoxic effects in sediments. Therefore, it allows a better assessment of the overall-toxicity in whole sediments. Recommendations and Outlook  The sediment contact test withMyriophyllum aquaticum is a valuable tool for the evaluation of the ecotoxicological risk potential of waters and sediments. It should become a complement to a standardised test battery generally used for the assessment of sediment toxicity.     

Contamination by PAHs,PCBs, PCPs and heavy metals in the mecoácfin lake estuarine water and sediments after oil spilling

Intention, Goal, Scope, Background  Environmental pollution caused by oil spills is a major ecological problem. Oil contamination in the environment is primarily evaluated by measuring the chemical concentrations of hydrocarbons. The results of chemical analyses are important for estimating water and sediment quality in the risk assessment to the flora and fauna of oil-contaminated sites. In the world there are lake ecosystems under permanent chemical stress due to urbanization and the oil industry. Studies, however, have been generally limited to petroleum compounds and have not considered other pollutants of the site like PCBs, polychlorinated pesticides and heavy metals. Objective  Water and sediment from stations in the Mecoacán Lake in the Mexican State of Tabasco were analyzed for polycy-clic aromatic compounds (PAHs), aliphatic hydrocarbons (AHs), polychlorinated biphenyls (PCBs), polychlorinated pesticides (PCPs) and heavy metals. The objective of this study was to examine the contaminant levels of the samples collected in February 1993 and 1996 after oil spills at the Mecoacán petroleum region. The goals of this study were to reveal the effect of the spills on the distribution of the hydrocarbons and assess the toxi-cological significance of the levels found. In addition, our aim was to examine the distribution of the PAHs in sediments from Mecoacán originated from both pyrolytic and petrogenic sources. Methods  Samples were collected from 19 stations and prepared according to the CARIPOL (Caribbean Pollution) methodology of the United Nations Environmental Programme (1992) of the Great Caribbean Region for hydrocarbons in marine and coastal water, and sediments. The gas-chromatographic and atomic absorption analysis of the samples was performed after sampling. Results and Discussion  Concentrations of PAHs in water ranged from 0.2 to 0.8 μg/l in 1993 and from 0.3 to 2.8 μg/l in 1996. The concentration of the 16 EPA-PAHs varied from 0.1 to 36 mg/kg dry weight in the lake sediment samples collected in 1996, while those of AHs, PCBs and PCPs ranged from 0.1 to 67 mg/kg, 0.1 to 59 μg/kg and 6 to 370 μg/kg, respectively. The most abundant contaminants in water were benz[a]anthracene and pyrene, in 1993 and 1996, respectively; while in sediments collected in 1996: Pyrene, C₂₄, 2-chlorobiphenyl and endrin predominated. Heavy metals (Cu, Pb and Zn) were found at low concentrations. Benzo[a]pyrene was detected in some sediment samples in varying amounts (0.2 to 0.3 mg/kg). Conclusions  The maximum total PAH concentration in sediments was found at sites near the oil fields and the AH concentration near the urban zone. The mayor pollutants in sediments were PAHs and AHs, and taking into regard the detected PAHs near the oil fields, the source was the oil spilling. The mean total PAH value in Mecoacan sediments of 6.4 mg/kg did not exceed the median range effects value (ERM) for total PAHs of 44.8 mg/kg. The measured organochlorine compounds and heavy metals were present in amounts much inferior to the ERM values. This study confirms that contaminants concentration in sediments did not exceed the environmental quality guideline for the 50% probability and no adverse effect can be expected. Recommendation and Outlook  Analysis revealed no indication of a contribution of PAHs, PCBs, PCPs and heavy metals to acute sediment toxicity. The results of this study demonstrate the importance of continuous monitoring of ecosystems exposed to pollution to make pre-spill data available in order to evaluate the real consequence of the spilling and its effect on flora and fauna.     

Determining tolerance limits for restoration and phytoremediation with Spartina patens in crude oil-contaminated sediment in greenhouse

Oil spills may detrimentally damage sensitive coastal habitats, such as coastal wetlands. Successful restoration of oiled habitats primarily depends on the tolerance of vegetation transplants to oil. In this study, tolerance limits of the dominant coastal brackish marsh plant Spartina patens to South Louisiana crude (SLC) oil and its phytoremediation effectiveness on petroleum hydrocarbons in wetland sediments were investigated in the greenhouse environment. Spartina patens was transplanted into brackish marsh sediments contaminated with SLC oil at concentrations of 0, 40, 80, 160, 320, 640 and 800 mg SLC oil g^?1 dry sediment. High oil concentrations adversely affected plant stem density, aboveground biomass and belowground biomass even one year after transplantation. At the 320 mg g^?1 oil dosage, plant belowground biomass was significantly lower than the control although aboveground variables were not significantly different from the control. All plant parameters mentioned above at the 640 mg g^?1 oil dosage were less than 50% of the control. Spartina patens did not survive the 800 mg g^?1 oil dosage. The tolerance limit of S. patens to SLC was estimated about 320 mg oil g^?1 dry sediment. In addition, S. patens transplants enhanced oil degradation in the sediment; concentrations of residual total petroleum hydrocarbons (TPH) in the sediments vegetated by S. patens were significantly lower than those of un-vegetated sediments for both the surface and subsurface sediment at the 40 and 160 mg g^?1 SLC oil dosages. Decreases in the concentrations of polycyclic aromatic hydrocarbons (PAHs) further demonstrated the capacity of S. patens to phytoremediate residual oil; residual total targeted PAHs in the phytoremediation treatment were less than 20% of the un-vegetated treatment at the 40 mg g^?1 oil dosage. These results demonstrated the potential of phytoremediation with S. patens to simultaneously restore and remediate petroleum-contaminated coastal marsh habitats.     

Potential effects of food addition to sediment on test conditions in sediment toxicity tests

Purpose  

Standardized sediment toxicity assays often employ periodic additions of uncontaminated food to sustain energy and growth requirements of the test organisms. Consequently, selective feeding on this uncontaminated food may reduce exposure to sediment particles containing the test substance. To address this issue, some standard guidelines propose to add food to the sediment before spiking with the test substance to account for multiple exposure routes, including ingestion of contaminated food. The present study focused on the influence of different feeding regimens and compositions of the aqueous medium on water quality (ammonia concentrations) and test organism development.