Applicability of the bioluminescence inhibition test in the 96-well microplate format for PAH-solutions and elutriates of PAH-contaminated soils

The use of conventional plastic microplates for a miniaturised luminescent bacteria test may result in an underestimation of the toxicity for poorly water soluble highly adsorbing toxicants such as PAHs. In this study, the suitability of microplates for testing elutriates of PAH-contaminated soils was investigated. The LUMIStox test was performed as the standard test in the miniaturised format using contaminated soil elutriates and aqueous solutions of four selected PAHs (viz. naphthalene (NAP), acenaphthene (ACE), fluorene (FLU), and phenanthrene (PHE)). For the aqueous PAH-solutions, we observed reduced light inhibition values for the miniaturised bioassay when using black microplates made of polypropylene (PP) and polystyrene (PS) compared to the standard LUMIStox test. This phenomenon was most likely due to adsorption of toxicants to the microplate surfaces with PAHs of lower water solubility being significantly more affected; however, after minimizing the exposure of samples to plastic surfaces, polystyrene microplates revealed equivalent performance (>80% 'relative' light inhibition) to the standard glass cuvette test system. For soil elutriates, black microplates again exhibited slightly lower light inhibition values while white plates made of PS and Barex resulted in a pronounced overestimation of toxicity for a coloured soil elutriate. In general, microplates were applicable for testing elutriates of PAH-contaminated soils. In cases where samples are coloured or turbid, the application of black microplates is recommended.     

Persistence of carbofuran in marine sand and water

Marine sand and seawater samples were collected in March 2002 from Laysan Island in the Hawaiian Islands National Wildlife Refuge, where a small area was contaminated by the carbamate insecticide carbofuran. Carbofuran was still detected at microg g(-1) levels in the Laysan sand after its identification in 1998 and initial observation of the toxicity in 1988. The persistence of carbofuran in the marine sand was investigated in the dark in a 30 degrees C oven, and in distilled deionized water and seawater samples exposed to artificial 300 nm light and to direct sunlight. The laboratory study showed a half-life (t1/2) of approximately 40 days for carbofuran in the native sand and in Ottawa sand. The photolysis of carbofuran was faster in seawater than in distilled deionized water when it was exposed to 300 nm light (t1/2, 0.1 vs. 3.1 h) and to direct sunlight (t1/2, 7.5 vs. 41.6 h). The large difference between the laboratory results and the field observation of carbofuran dissipation suggests that carbofuran degradation at the remote, undisturbed marine site may be governed by its unique environmental factors.     

Urease inhibition: a tool for toxicity identification in sediment elutriates

A method for the detection and confirmation of heavy metal toxicants in sediment elutriates based on a urease inhibition assay, ICP-AES analysis and EDTA chelation in the frame of toxicity identification evaluation (TIE) is presented. Zinc was identified as the major toxicant in pHstat elutriates of sediments of the river Saale (Germany). Implications of natural and anthropogenic chelating agents, which are frequently present in environmental samples, on toxicity confirmation of heavy metals based on the toxic unit approach are discussed.     

Effects of anthracene, pyrene and benzo[a]pyrene spiking and sewage sludge compost amendment on soil ecotoxicity during a bioremediation process

The fate of spiked anthracene, pyrene and benzo[a]pyrene in soil with or without sewage sludge compost was assessed during a 6-month bioremediation process simulating landfarming. Bioassays and physico-chemical analyses were employed to monitor toxicity change in soil samples and elutriates through ten sampling campaigns. Pearson product-moment correlation coefficient was determined to measure the strength of relationship between bioassays and physico-chemical analyses. The PAH dissipation in soil was enhanced after the first water addition, and the remaining amounts at the end of the experiment were positively correlated to the number of benzene rings and the presence of sewage sludge compost. Toxicity of soil elutriates to Daphnia magna was evident at early stages, originating exclusively from sewage sludge compost amendment. The lettuce root elongation was continuously inhibited by elutriates for all the treatments including control soil, probably due to high salinity or to unaddressed leachable phytotoxic compounds that were present in the experimental soil. The newly developed direct solid-phase chronic toxicity test using ostracod (Heterocypris incongruens) succeeded in evaluating the soil-bound PAH toxicity, as PAHs could not be detected in elutriates.     

Influence of organic solvents on the sensitivity of a bioluminescence toxicity test with Vibrio harveyi

The possibility of using organic solvents (OSs) to increase the susceptibility of bioluminescent microorganisms in a bioassay for assessing the toxicity of chemicals dissolved in water was investigated. To conduct the tests acetonitrile, dimethyl sulfoxide, ethanol, methanol, and isopropanol were used as OSs and Cd, Hg, and Zn as reference toxicants. The addition of OSs modified the toxicity of the three metals to Vibrio harveyi, according on the bioluminescence assay used. The sensitivity of the luminescence bioassay for Hg increased in the presence of the five OSs, thus indicating a greater toxic effect. However, the sensitivity of the assay for the other two metals, Cd and Zn, increased or decreased (lesser toxic effect) depending on the concentration at which the OSs were used. No correlation was observed between the concentration of the five OSs and the toxicity of the three reference toxicants. From this it can be deduced that none of these OSs could be recommended for increasing generically the sensitivity of toxicity biotests using V. harveyi.     

Integrative assessment of marine pollution in Galician estuaries using sediment chemistry,mussel bioaccumulation,and embryo-larval toxicity bioassays

An integrative assessment of environmental quality was carried out in selected sites along the Galician coast (NW Iberian Peninsula) combining analytical chemistry of seawater and sediments, bioaccumulation in the marine mussel, and embryo-larval sediment toxicity bioassays, in order to link biological and chemical criteria for the assessment of coastal pollution. Maximum values of Hg and Cu in seawater, sediment and mussels, were found in the inner part of Ria of Pontevedra, while maximum levels of organics (polychlorinated biphenyls, hexachlorobenzene and aldrin) were found in mussels from A Coru?a. Outstanding values of Cu, Pb and Zn have been found in seawater and sediment from a single site, P3, which also was the most toxic in the embryo-larval bioassays performed with four different phyla of marine organisms: mollusks, echinoderms, arthropods and chordates. Sediment quality effects range-median values provided a valuable reference to predict biological effects from sediment chemistry data, while effects range-low values were too conservative. Sediment toxicity could also be predicted by using a toxic-unit model based on published EC50 values for trace metals and mobilization factors independently obtained from measurements of metal contents in sediments and their elutriates. When chemical and toxicological data are independently used to arrange sampling sites by using non-metric multidimensional scaling, a remarkable degree of concordance between both types of configurations could be observed.     

Effects of storage method and duration on the toxicity of marine sediments to embryos of Crassostrea gigas oysters

The objective of laboratory sediment bioassays is to estimate in situ toxicity. This goal is difficult to achieve, as one of the main limitations of sediment toxicity tests is disruption of sediment geochemistry during sampling, handling and preservation. The effects of storage on the estimation of marine sediment toxicity to Crassostrea gigas embryos and larvae were investigated. Three storage methods and four storage periods were compared with three different sediment types contaminated by heavy metals, polycyclic aromatic hydrocarbons (PAHs) and both contaminants. Freezing and freeze-drying considerably increased the toxicity of decanted sediments and their elutriates as compared to the toxicity obtained with fresh sediments. Concerning the elutriates, the toxicity found with frozen and freeze-dried sediments was correlated with DOC, ammonia and PAH contents. However, the toxicity of fresh sediments kept at 4 degrees C increased with increasing duration of storage and was also correlated with the amount of ammonia in the elutriates.     

The toxicity and concentrations of PAHs in creosote-contaminated lake sediment

Sediment samples, divided into three layers (0-10, 10-20 and 20-30 cm), were collected from 16 sites in Lake J?ms?nvesi, Central Finland. The acute toxicity of pore waters and elutriates (sediment + water 1:4 v/v) were studied by bioluminescence inhibition test and by immobilisation of water fleas (Daphnia magna Straus). Concentrations of polycyclic aromatic hydrocarbons (PAHs) in sediments and elutriates were measured by gas chromatography using flame ionization detection (GC/FID). The highest total PAH concentration was 3.3 mg/g dry weight in the sediment and up to 1.7 mg/l in the elutriate of the uppermost (0-10 cm) layer, also being the most toxic to photoluminencent bacteria and water flea. When sediment and water mix, like during dredging operations, toxic compounds may spread from the sediment to the water column and can pose on environmental risk.     

Toxicity and heavy metal contamination of surficial sediments from the Bay of Thessaloniki (Northwestern Aegean Sea) Greece

Surficial sediments (the fraction < 2000 microm) from the Bay of Thessaloniki, in the Northwestern Aegean Sea, Greece were examined for heavy metal and organic carbon contents, as well as for acute LUMIStox toxicity of pore waters (PWs), wet sediment elutriates (WSEs) and dry sediment elutriates (DSEs) obtained from the solid material remaining after PW extraction. WSEs where not toxic. EC20/50 values could be measured in some DSE and PW samples. In all sediment samples, the DSE toxicity was greater than the corresponding PW toxicity. Sediment concentrations of total and labile Cd, Pb, Cu, Cr, Zn, Mn, Ni and Fe were determined and evaluated in relation to sediment quality guidelines. Pollution levels ranged from low to high for certain metals. A misfit between sediment toxicities calculated from heavy metal concentrations and those biologically measured was observed. Toxicity values were in general poorly correlated with sediment's heavy metal or organic carbon content. As significant correlation was however found between the DSE toxicity with total Ni and labile Fe, as well as for the PW toxicity with total Ni, labile Fe and labile Cr.     

Potential impacts of water injection dredging on water quality and ecotoxicity in Limehouse Basin, River Thames, SE England, UK

The use of water injection dredging (WID) is increasing in the UK's inland waterways and marinas. Jets of water are injected under low pressure directly into bottom sediment creating a turbulent water-sediment mixture that flows under the influence of gravity. Many of these sediments are highly contaminated and little is known of the effects of contaminant release on water quality or the risk to biota living in both the sediment and the water column. Sediment cores were collected from Limehouse Basin, a proposed WID site in SE England and current sediment toxicity was assessed using a number of techniques. Comparison of metal data to US sediment quality guidelines indicated intermediate levels of toxicity while, calculation of acid volatile sulphide to simultaneously extracted metal ratios underestimated the potential toxicity to sediment dwelling organisms. In contrast, porewater ammonia concentrations were in excess of all published ecotoxicological guidelines and indicate serious risk to biota. Re-suspension experiments were used to mimic the effects of WID on overlying water quality and ecotoxicity tests were carried out on elutriates using Daphnia magna to examine the impacts on biota. Concentrations of a range of metals in the elutriates predict that adverse biological effects would be observed during WID, however only 10% of the elutriate samples caused an adverse effect on Daphnia. Limehouse Basin is a complex aquatic environment receiving predominantly fresh waters while the sediments have high porewater chloride concentrations reminiscent of previous tidal inputs to the basin, making the choice of test organism problematic.     

发光细菌法在水质综合毒性在线检测中的应用

以海洋发光细菌费氏弧菌（Vibriofischeri）作为检测生物，采用冻干菌粉快速复苏技术，研究费氏弧菌在水质检测中的最佳测试温度和有效性测试条件，并对硫酸锌等多种毒物和几种实际水样进行发光抑制作用分析。研究表明，费氏弧菌冻干粉复苏菌液保存在2～5℃条件下能有效测试7d，最佳测试温度为15℃，最佳测试时间为15min。氯化汞、硫酸锌、硫酸镉等重金属和苯胺、多菌灵、甲醛等有机毒物对费氏弧菌均具有较强的光抑制作用，也即费氏弧菌对以上毒物较为敏感，并能够连续7d保持对同一浓度硫酸锌的敏感性较为一致。对几种实际水样的测试和分析表明，以费氏弧菌为指示生物的发光细菌法能够应用于水质环境安全的综合毒性在线监测预警中。     

Evaluation of the toxicity of two soils from Jales Mine (Portugal) using aquatic bioassays

Soil contamination can be one path for streams and groundwater contamination. As a complement of chemical analysis and total contaminants determination, bioassays can provide information on the bioavailable fraction of chemical compounds, focusing on the retention and habitat function of soils. In this study the evaluation of the toxicity of two soils from the abandoned Jales Mine (Portugal) regarded both functions. The buffer capacity of soils was tested with bioassays carried out using the cladoceran Daphnia magna and the marine bacteria Vibrio fischeri. The habitat function of soils was evaluated with the reproduction bioassay with the collembolan Folsomia candida. The Microtox solid-phase test was performed with V. fischeri using soil as test medium, and soil elutriates were extracted to perform the Microtox basic test, and an immobilization and reproduction bioassay with D. magna. The marine bacteria showed high sensitivity to the soil with low heavy metal content (JNC soil) and to JNC soil elutriates, while the soil with highest heavy metal content (JC soil) or soil elutriates exposure did not cause any toxic effect. In the bioassays with D. magna, organisms showed sensitivity to JNC and also to JC soil elutriates. Both mobilization and reproduction features were inhibited. The bioassay with F. candida did not reflect any influence of the contaminants on their reproduction. Although JNC soil presented lower heavy metal contents, elutriates showed different patterns of contamination when compared to JC soil and elutriates, which indicates different retention and buffer capacities between soils. Results obtained in this study underlined the sensitivity and importance of soil elutriate bioassays with aquatic organisms in the evaluation strategy in soil ERA processes.     

Comparison of test procedures for sediment toxicity evaluation with Vibrio fischeri bacteria

The use of bacterial luminescence assays is particularly effective in the contaminated sediment evaluation. In the present study, a comparison was made of different baterial luminescence assays including acute toxicity of elutriates with Microtox® and LUMIStox®; chronic toxicity of elutriates with LUMIStox®; and, acute toxicity of solvent extracts with both the tests and the Solid Phase test with Microtox. The toxicity assay procedures were utilised for the Po River sediment toxicity evaluation, testing samples with different physical and chemical characteristics. The results evidentiated that each sediment test procedures provided indipendent and complementary ecotoxicological responces usefull for a sediment classification of the most polluted samples.     

Site-specific water quality guidelines: 1. Derivation approaches based on physicochemical, ecotoxicological and ecological data

Generic water quality guidelines (WQGs) are developed by countries/regions as broad scale tools to assist with the protection of aquatic ecosystems from the impacts of toxicants. However, since generic WQGs cannot adequately account for the many environmental factors that may affect toxicity at a particular site, site-specific WQGs are often needed, especially for high environmental value ecosystems. The Australian and New Zealand Guidelines for Fresh and Marine Water Quality provide comprehensive guidance on methods for refining or deriving WQGs for site-specific purposes. This paper describes three such methods for deriving site-specific WQGs, namely: (1) using local reference water quality data, (2) using biological effects data from laboratory-based toxicity testing, and (3) using biological effects data from field surveys. Two case studies related to the assessment of impacts arising from mining operations in northern Australia are used to illustrate the application of these methods. Finally, the potential of several emerging methods designed to assess thresholds of ecological change from field data for deriving site-specific WQGs is discussed. Ideally, multiple lines of evidence approaches, integrating both laboratory and field data, are recommended for deriving site-specific WQGs.     

Microscale Toxicity Identification Evaluation (TIE) for interstitial water of estuarine sediments affected by multiple sources of pollution

Estuaries in the world are affected by different contamination sources related to urbanisation and port/industrial activities. Identifying the substances responsible for the environmental toxicity in estuaries is challenging due to the multitude of stressors, both natural and anthropogenic. The Toxicity Identification and Evaluation (TIE) is a suitable way of determining causes of toxicity of sediments, but it poses difficulties since its application is labour intensive and time consuming. The aim of this study is to evaluate the diagnosis provided by a TIE based on microscale embryotoxicity tests with interstitial water (IW) to identify toxicants in estuarine sediments affected by multiple stressors. TIE showed toxicity due to different combinations of metals, apolar organic compounds, ammonia and sulphides, depending on the contamination source closest to the sampling station. The microscale TIE was able to discern different toxicants on sites subject to different contamination sources. There is good agreement between the results indicated in the TIE and the chemical analyses in whole sediment, although there are some disagreements, either due to the sensitivity of the test used, or due to the particularities of the use of interstitial water to assess the sediment toxicity. The improvement of TIE methods focused on identifying toxicants in multiple-stressed estuarine areas are crucial to discern contamination sources and subsidise management strategies.

     

Species-specific responses to metals in organically enriched river water, with emphasis on effects of humic acids

Invertebrate communities in polluted rivers are often exposed to a wide variety of compounds. Due to complex interactions, 'pollution tolerant' species are not necessarily the most tolerant species to toxicants tested under standard laboratory conditions. It was hypothesized that the distribution of species in polluted rivers is not only dependent on the tolerance of species to toxicants, but also on species-specific capacities to modify or compensate for negative effects of toxicants. To test this hypothesis, species-specific responses to metals in organically enriched river water were studied under controlled conditions. The zebra mussel Dreissena polymorpha and the midge Chironomus riparius were exposed to metal-polluted water from the River Dommel. Additionally, the (interactive) effects of metals and humic acids (HA) on both species were evaluated. In spite of a lower tolerance of Chironomus riparius to metals in laboratory studies, the midge was the most tolerant of the two test species to metal-polluted site water. The results indicated that the sensitivities of the two test species determined in laboratory tests were inversely related to their sensitivities to polluted river water. In accordance with these results, midge larvae were protected from copper (Cu) toxicity by HA, while metal toxicity was not reduced (Cu) or even amplified (cadmium) by HA for the zebra mussel. Thus, the presence of (naturally occurring) HA in site water may partly account for discrepancies between responses of species to bioassays and toxicity tests. It is suggested that these differences in responses to metals in site water are strongly influenced by species-specific preferences for organic compounds (like HA). It is concluded that the response to organic compounds present in site water largely determines whether a species is classified as 'pollution tolerant' or 'pollution sensitive'.     

Study of chlorothalonil photodegradation in natural waters and in the presence of humic substances

Photodegradation of chlorothalonil was studied in different natural waters (sea, river and lake) as well as in distilled water under natural and simulated solar irradiation. The effect of dissolved organic matter (DOM) such as humic and fulvic substances on the photodegradation rate of chlorothalonil was also studied in simulated sunlight. The presence of DOM enhanced the photodegradation of chlorothalonil with the exception of seawater. The kinetics were determined through gas chromatography electron capture detection (GC/ECD) and the photodegradation proceeds via pseudo-first-order reaction in all cases. Half-life ranged from 1 to 48 h. In natural and humic water chlorothalonil photodegradation gave rise to two different intermediates compared to distilled water demonstrating that the transformation of chlorothalonil depend on the constitution of the irradiated media and especially from DOM. The byproducts identified by GC/MS techniques were: chloro-1,3-dicyanobenzene, dichloro-1,3-dicyanobenzene, trichloro-1,3-dicyanobenzene and benzamide.     

Compatibility of hydroxypropyl-beta-cyclodextrin with algal toxicity bioassays

Numerous reports have indicated that hydrophobic organic compound bioaccessibility in sediment and soil can be determined by extraction using aqueous hydroxypropyl-β-cyclodextrin (HPCD) solutions. This study establishes the compatibility of HPCD with Selenastrum capricornutum and assesses whether its presence influences the toxicity of reference toxicants. Algal growth inhibition (72 h) showed no significant (P > 0.05) difference at HPCD concentrations up to and including 20 mM. HPCD presence did not influence the toxicity of the inorganic reference toxicant (ZnSO₄), with IC50 values of 0.82 μM and 0.85 μM, in the presence and absence of HPCD (20 mM), respectively. However, HPCD presence (20 mM) reduced the toxicity of 2,4-dichlorophenol and the herbicides diuron and isoproturon. These reductions were attributed to inclusion complex formation between the toxicants and the HPCD cavity. Liberation of complexed toxicants, by sample manipulation prior to toxicity assessment, is proposed to provide a sensitive, high throughput, bioassay that reflects compound bioaccessibility.     

Spore release by the green alga Ulva: a quantitative assay to evaluate aquatic toxicants

A toxicity test using spore release of the aquatic green alga, Ulva, was developed and evaluated by assessing the toxicity of different organic and inorganic chemicals and elutriates of sewage or waste sludge. The toxic ranking of four metals was: Cu (EC50 of 0.040mgL(-1))>Cd (0.095mgL(-1))>Pb (0.489mgL(-1))>Zn (0.572mgL(-1)). The EC50 for TBTO ranged from 24 to 63microgL(-1). The most toxic VOC was formalin (EC50 of 0.788microlL(-1)) and the least toxic was acetone. Spore release was significantly inhibited in all elutriates; the greatest and least toxic effects were for industrial sewage (3.29%) and filtration bed (10.08%), respectively. The bioassay is simple, inexpensive and sensitive. The cosmopolitan distribution of Ulva means that the test would have a potential application worldwide.     

Complexation of arsenate with humic substance in water extract of compost

The interactions of environmental toxicants with organic substances affect the speciation and dynamics, and subsequent toxicity, mobility, and fate of toxicants in the environment. For the purpose of understanding the complexation of As(V) with humic substances, arsenate-containing solutions with As concentrations from 1 to 8 mg l⁻¹ were prepared to react with the water extract of compost (WEC). All the reaction systems including the control were incubated for 48 h at 25 °C. The complexation of As(V) with humic substance was examined by dialysis and ion exchange techniques. From 30% to 51% of added As(V) reacted with organic substance in WEC to form an As–metal–organic complex. This was verified as a hydrophobic organic fraction after separation of As–metal–organic complex fraction from the hydrophilic fraction by XAD-8 resin. The complex substance was also identified as a humic substance by the method of proton binding formation function determination. This suggests that cations, such as Ca and Mg, and especially Fe, Al, and Mn act in cation bridging in the complexation of As(V) with humic substance. The role of metals in the complexation of As(V) with humic substance in terrestrial and especially aquatic environments thus merits close attention.