Battery Of Bioassays Research Articles

Assessing Domestic Wastewater Effluent with a Battery of Bioassays after Treatment with a Specific Consortium of Microalgae and Different Flocculation Methods

Wastewater stabilization ponds (WSP) are a common method to treat wastewater before release. The purpose of the current study was to find the most inexpensive coagulation and flocculation reagents to use during harvesting of WSP algae from effluent so that water without algae can be generated for irrigation purposes. In the study, the toxicity of effluent from the algae-based WSP system after treatment with alum and chitosan was investigated. The chemical composition of the resultant supernatants was tested in conjunction with a battery of bioassays (Daphnia magna, Allium cepa, Lactuca sativa and Triticum aestivum). The bioindicator organisms were exposed to varying concentrations of the supernatant resulting from harvesting the algae biomass with chitosan (0.368 g/L chitosan) and aluminium sulphate (9.96 g/L powder). Alum was lethal to D. magna, while chitosan did not seem to affect the Daphnia test specimens. Undiluted chitosan supernatant and 100% alum resulted in minimal inhibition of the rate of T. aestivum seed germination, while both resulted in significant inhibition of the L. sativa seed germination rates. Similarly, minimal effects were found for the root growth in T. aestivum, with significant root growth inhibition observed in L. sativa and A. cepa. In A. cepa, clear differences were observed in the inhibition of root growth when the two reagents were compared. The alum and chitosan supernatants had a total carbon, iron and sulphate concentration of 327.78 mg/L, 0.12 mg/L and 68.44 mg/L and 182.04 mg/L, 0.16 mg/L and 166.80 mg/L, respectively. Based on the test specimens’ response, chitosan as flocculation method was less toxic when compared with alum coagulation, even though both algae harvesting methods changed the chemical composition of the algae-treated water.

Integrated characterization and risk management of marine sediments: The case study of the industrialized Bagnoli area (Naples, Italy)

The aim of the present work is to demonstrate the practical importance of a multidisciplinary approach and weighted criteria to synthesize and integrate different typologies of data (or lines of evidence, LOEs), including chemical levels in marine sediments, their bioavailability to specific indicator species, ecotoxicological effects measured through subcellular biomarkers and batteries of bioassays, and potential impacts of pollution on local benthic communities. The area of Bagnoli (Gulf of Naples, Southern Italy) was selected as a model case-study, as it is a coastal area chronically impacted by massive industrial contamination (trace metals and hydrocarbons), and dismissed decades ago without any subsequent remediation or habitat restoration. The results of each LOE were elaborated to provide specific hazard indices before their overall integration in a weight of evidence (WOE) evaluation. Levels of some trace metals and PAHs revealed a severe contamination in the entire study area. Bioavailability of hydrocarbons was evident particularly for high molecular weight PAHs, which also caused significant variations of cellular biomarkers, such as cytochrome P450 metabolization in fish, lysosomal membrane destabilization in mussels, genotoxic effects both in fish and molluscs. The results of a battery of bioassays indicated less marked responses compared to those obtained from chemical and biomarkers analyses, with acute toxicity still present in sediments close to the source of contamination. The analysis of benthic assemblages showed limited evidence of impact in the whole area, indicating a good functioning of local ecosystems at chronic contamination. Overall, the results of this study confirm the need of combining chemical and biological data, the quantitative characterization of various typologies of hazard and the importance of assessing an integrated environmental WOE risk, to orientate specific and scientifically-supported management options in industrialized areas.

Assessment of hazardous property HP 14 using ecotoxicological tests: a case study of weathered coal fly ash.

The classification of wastes regarding hazardous property HP 14 (ecotoxicity) is essential for proper waste management. In the EU, HP 14 has been estimated based on waste chemical composition rather than using biotests, and guidelines for experimental assessment are still lacking. This study aims at evaluating the potential ecotoxicological impacts of weathered coal fly ash (CFA) from a landfill, as a case study to assess the current EU methodology used to classify wastes regarding HP 14. A large amount of CFA is still landfilled, but its valorisation would be of interest. The analysis was based on the chemical composition of CFA (in ClassifyMyWaste software), and on a battery of five biotests applied to eluates, with Lepidium sativum, Aliivibrio fischeri, Raphidocelis subcapitata, Lemna minor and Daphnia magna. Through chemical analysis, most of the simulations with data from the literature indicated "Possible Hazard", including the sample of this work. Biotests revealed low impairment for most endpoints. D. magna was the most sensitive organism, but the inhibitory effect was significantly reduced after pH adjustment of the eluate. The test with A. fischeri does not seem to be adequate to assess CFA due to the high variability observed in results. The methodology involving a simple battery of bioassays was proven to be enlightening, providing relevant results for HP 14 assessment. The chosen battery of biotests (excluding the A. fischeri test) may be a good starting point to represent the aquatic environment in this context. In short, it seems that weathered CFA can be considered non-hazardous, and therefore the material under analysis could be valorised in practical applications without significant ecotoxic effect on the environment.

Ecotoxicity and antibiotic resistance of wastewater during transport in an urban sewage network.

Urban wastewater (UWW) management usually entails biological and physicochemical monitoring due to its potential impact on the quality of the receiving environment. A major component of a sewage system is the pipe network leading the water to the treatment plant. Up to now, few studies have been conducted on the diverse phenomena that may affect the characteristics of the water during its transportation. In this study, ecotoxicity and potential antibiotic resistance were used in a global method to assess the change of UWW quality in a sewage system and determine if sewer pipes can act as a bioreactor spread. Three bioassays were conducted to assess the ecotoxicity of the samples and the concentration and relative abundance of two classes of integrons (as a proxy for antibiotic resistance) were measured. The results of the bioassay battery do not show a pattern, despite the fact that differences were noticeable between upstream and downstream samples. Antibiotic resistance appeared to decrease during transport in the pipe as the concentration and relative abundance of integrons decreased during several campaigns. This result should be confirmed in other sewer networks but already provides useful information for the management of urban sewage system.

Ecotoxicological test based on inhibition of fungal laccase activity: Application to agrochemicals and the monitoring of pesticide degradation processes.

Ecotoxicological evaluations require the use of assays with several bioindicators from different trophic levels. Only a few ecotoxicological tests using fungi have been developed, reason why, detection of adverse effects from compounds that exert fungicide action may be overlooked. This work developed a toxicity test based on the inhibition of laccase enzymatic activity in the fungus Trametes versicolor. The test was applied to several fungicides and succeeded to determine inhibition values (half maximum effective concentration, EC50) for most of them (flusilazole, imazalil, pyrimethanil, tetraconazole), though a clear dose-response was not evident for others (thiabendazole, metalaxyl). The application on atrazine (herbicide), imidacloprid (insecticide) and oxytetracycline (antibiotic), proved the proposed test is suitable towards other agrochemicals. The test was also used to estimate the detoxification resulting from two different approaches employed in the removal of agrochemicals. (a) First, in the liquid-phase elimination by fungal biomass simultaneously removing atrazine, imazalil, tebuconazole and triadimenol, the test showed a significant decrease in toxicity by biodegradation (adsorption contribution to detoxification was negligible). (b) Second, a solid-phase biomixture (used for pesticide degradation from agricultural wastewater) partially removed atrazine, imazalil, metalaxyl and pyrimethanil after 33d; nonetheless, this system could not reduce the toxicity of the matrix, and higher laccase inhibition was detected after the treatment. The design test increases the battery of available bioassays to determine the toxicity of agrochemicals, and provides an interesting tool to monitor biodegradation processes.

A methodological approach for ecotoxicological characterization of non-hazardous sediments for their beneficial reuse

In France, the beneficial reuse of sediments is conditioned upon a demonstration of environmental acceptability for each scenario. Such evidence has to be provided in application of the NF EN 12920 +A1 standard methodologies. Before the application of risk assessment methodology, the HP14 hazard property must be measured using specific bioassays. However, sediments may have ecotoxicological impacts despite a non-hazardous classification. The application of a battery of bioassays sensitive to non-hazardous sediments could allow to select sediments before choosing the beneficial reuse option. First, HP14 methodologies were applied on the studied sediments. These sediments had to be managed on shore according to French legislation. Then, following recommendations from a previous study, other ecotoxicological tests and modified ecotoxicological tests from HP14 methodology were selected. Ostracods mortality and growth tests were selected as a complement because its sensitivity to sediment ecotoxicity is known. To take into account the effects of extraction method and liquid/solid ratio on ecotoxicological impact of sediments, ecotoxicological tests have been applied to percolates. They have been obtained from up-flow percolation tests following the standard NF EN 14405. For each percolate, the rotifer clones reproduction test and the ostracod mortality and growth tests were performed. Contaminant analyses of sediments studied reveal differences in contamination between non-hazardous sediments. The majority of ecotoxicological tests carried out in the HP14 test batteries do not differentiate between non-hazardous sediments with low ecotoxicity. However, inhibition of rotifer reproduction and inhibition of plant germination and growth tests can be used. Among the complementary bioassays performed, the ostracods growth inhibition and multigenerational Daphnia reproduction inhibition tests are especially sensitive. Bioassays performed on percolates reveal low ecotoxicities that were not detected with conventional bioassays. For these tests, maximum ecotoxicities are observed on the first percolates for most of the tested sediments, and ecotoxicity effects decrease with the increase in the liquid/solid ratio. The results enable us to propose a bioassay battery able to highlight the intrinsic toxicity of non-hazardous sediments. This is achieved through bioassays and using up-flow percolation columns to detect low ecotoxicity. This battery can be integrated into the SEDIMATERIAUX approach to ensure the choice of the beneficial reuse option for such sediment. As some of the proposed bioassays remain unsuitable for marine sediments, future studies will adapt the methodology to the presence of marine salts.

Toxicity of PM2.5 Based on a Battery of Bioassays

To accurately evaluate the biotoxicity of PM2.5, PM2.5 samples from winter fog-haze days, winter normal days, and summer days in Changzhou were selected for evaluation based on the acute toxicity of luminescent bacteria and zebrafish embryos and the cytotoxicity of human lung adenocarcinoma cells in vitro (A549). The three atmospheric conditions and the physical and chemical indicators were also evaluated using correlation analysis. The PM2.5 samples showed either acute or developmental toxicity during all three periods. The toxicity unit (TU) of the luminescent bacteria for the winter fog-haze days, winter normal days, and summer days were 1.74 (toxic), 1.19 (toxic), and 0.92 (slightly toxic), respectively. The maximum TU of the zebrafish embryos was for winter normal days (TU=1.14, toxic) followed by winter fog-haze days (TU=0.79, slightly toxic), and summer days (TU=0, non-toxic). The highest TU of A549 was for winter fog-haze days (TU=0.61, toxic) followed by summer days (TU=0.38, toxic) and winter normal days (TU=0.31, toxic). With respect to developmental toxicity, with the exception of summer day samples, the PM2.5 samples from the other two periods had detrimental effects on the development of zebrafish embryos, mainly showing pericardial edema, a bent notochord, and tail deformity. The average toxicity (AvTx), toxic print (TxPr), and most sensitive test (MST) indices showed that the PM2.5 samples from winter fog-haze days and winter normal days exhibited toxicity, while samples from the summer days showed slight toxicity; PM2.5 samples from winter fog-haze days had the highest level of comprehensive toxicity. In addition, luminescent bacteria were the most sensitive to PM2.5 samples, followed by zebrafish embryos and A549. The results of chemical analysis and biological toxicity tests show that the pollutants contained in PM2.5 have a biological toxicity effect, which can provide a basis for the comprehensive assessment of PM2.5 biological toxicity and human health risks.

Exposure to the azo dye Direct blue 15 produces toxic effects on microalgae, cladocerans, and zebrafish embryos.

Aquatic pollution caused by dyes has increased together with the growth of activities using colorants such as the textile, leather, food, and agrochemicals industries. Because most popular azo dyes are synthesized from benzidine, a carcinogenic compound, a threat to aquatic biota could be expected. The use of single species for toxicity assessment provides limited data, so a battery of test organisms, including representatives of different trophic levels such as algae, zooplankters, and fish, could undoubtedly provide more information. Therefore, our study was aimed at evaluating the toxic effect of the azo dye Direct blue 15 (DB15) on a battery of bioassays using a primary producer (Pseudokirchneriella subcapitata), a primary consumer (Ceriodaphnia dubia), and a secondary consumer (Danio rerio). P. subcapitata was more sensitive to DB15 (IC50 = 15.99 mg L-1) than C. dubia (LC50: 450 mg L-1). In the algae exposed to DB15, chlorophyll-a and -b were significantly increased, and carotenoids were reduced. The concentrations of protein, carbohydrates, and lipids per cell in P. subcapitata exposed to all DB15 concentrations were significantly higher than that measured in control. At 25 mg L-1 of DB15, survival, total progeny, and the number of released clutches were significantly decreased, and the start of reproduction was delayed in C. dubia. DB15 did not induce lethal or sublethal effects in D. rerio embryos at any of the tested concentrations from 24 to 72 h post-fertilization (hpf), but from 96 to 144 hpf, the larvae exposed to 100 and 500 mg L-1 developed yolk sac edema, curved tail, and skeletal deformations. After 144 hpf, DB15 produced a significant increase in embryos without a heartbeat, as the concentration of dye raised. The textile-used, azo dye DB15, caused toxic effects of different magnitude on microalgae, cladocerans, and zebrafish embryos; for this reason, the discharge of this colorant into waterbodies should be regulated to prevent environmental impacts.

Application of a Weight of Evidence Approach for Monitoring Complex Environmental Scenarios: the Case-Study of Off-Shore Platforms

Multidisciplinary investigations based on integration of chemical and biological measurements, represent an added value to monitoring and management protocols, and their use is recommended by European Directives to evaluate environmental status of aquatic ecosystems. Nonetheless, assessing the overall significance of results from different studies is still a challenge. This work was aimed to present a quantitative Weight Of Evidence (WOE) model (Sediqualsoft) to integrate huge amounts of heterogeneous data and to validate this approach in complex monitoring scenarios. Using the case-study of off-shore platforms in the Adriatic Sea, procedures are presented to elaborate different typologies of data (lines of evidence, LOEs), including chemical characterization of sediments, bioavailability, biomarkers, ecotoxicological bioassays and benthic communities. These data are initially evaluated by logical flowcharts and mathematical algorithms, which provide specific hazard indices for each considered LOE, before their weighting and integration in an environmental risk index. The monitoring study selected for the WOE elaboration consisted on chemical analyses of trace metals, aliphatic hydrocarbons, polycyclic aromatic hydrocarbons carried out on 60 sediment samples; the same samples were characterized for the status of benthic communities; bioavailability of metals from sediments was assessed at laboratory conditions on the polychaete Hediste diversicolor, while bioaccumulation of inorganic and organic chemicals and biomarkers were measured in native and transplanted mussels; ecotoxicological properties of sediments were evaluated through a battery of bioassays determining algal growth of the diatom Phaeodactylum tricornutum, bioluminescence of the marine bacterium Vibrio fischeri, survival of the copepod Acartia tonsa and embryotoxicity of sea urchin Paracentrotus lividus. Almost 7000 analytical results were elaborated and summarized in specific hazard indices. The WOE integration of multiple typologies of data allowed more robust and weighted conclusions compared to the use of individual LOEs, highlighting the feasibility of this procedure for multidisciplinary monitoring and risk assessment approaches. On a practical side, the WOE evidences also suggested a revision of actual monitoring procedures. The proposed WOE model appears a useful tool to summarize large datasets of complex data in integrative indices, and to simplify the interpretation for stakeholders and decision makers, thus supporting a more comprehensive process of “site-oriented” management decisions.

Toxicity evaluation of olive oil mill wastewater and its polar fraction using multiple whole-organism bioassays

Olive mill wastewater (OMW) as a by-product of olive oil extraction process has significant polluting properties mainly related to high organic load, increased COD/BOD ratio, high phenolic content and relatively acidic pH. Raw OMW from Slovenian Istria olive oil mill and its polar fraction were investigated in this study. Chemical characterization of OMW polar fraction identified tyrosol as the most abundant phenolic product, followed by catechol. Lethal and sub-lethal effects of OMW matrix and its polar fraction were tested using a battery of bioassays with model organisms: bacteria Vibrio fischeri, algae Chlorella vulgaris, water fleas Daphnia magna, zebrafish Danio rerio embryos, clover Trifolium repens and wheat Triticum aestivum. Raw OMW sample was the most toxic to V. fischeri (EC50 = 0.24% of OMW sample final concentration), followed by D. magna (EC50 = 1.43%), C. vulgaris (EC50 = 5.20%), D. rerio (EC50 = 7.05%), seeds T. repens (EC50 = 8.68%) and T. aestivum (EC50 = 11.58%). Similar toxicity trend was observed during exposure to OMW polar fraction, showing EC50 values 2.75–4.11 times lower comparing to raw OMW. Tested samples induced also sub-acute effects to clover and wheat (decreased roots, sprouts elongation); and to zebrafish embryos (increased mortality, higher abnormality rate, decreased hatching and pigmentation formation rate). A comprehensive approach using a battery of bioassays, like those used in this study should be applied during ecotoxicity monitoring of untreated and treated OMW.

Analytical methodology for evaluation of the effects of different leachate treatment methods

Characterisation of the efficiency of leachate treatment methods when considering differentgroups of pollutants as phenolic compounds, PCB,s, PBDEs and phthalates is discussed.Strategies for measurements, identification and quantifications of the organic pollutants arepresented and the use of supported liquid membrane extraction (SLM) and micro porousmembrane extraction (MMLLE) in particular, where small and medium molecules areseparated from large molecules. Acute and chronic toxicity tests by biological methods andthe choice of organisms are also presented as well as the LAQUA-protocol used at the pilotplant in Kristianstad, Sweden. In acute toxicity tests of the leachate a clear accommodating onthe immobility of the Artemia larvae and increasing concentration of leachate was noted.Different species react different on different compounds and therefore on complex effluentsas landfill leachates the use of a battery of bioassays is recommended.

Effect-based nationwide surface water quality assessment to identify ecotoxicological risks

A large portion of the toxic effects observed in surface waters cannot be attributed to compounds regularly measured by water authorities. Hence, there is an urgent need for an effect-based monitoring strategy that employs bioassays to identify environmental risks. The aim of the present study was to perform an effect-based nationwide water quality assessment to identify ecotoxicological risks in a wide variety of surface waters. At 45 locations silicone rubbers and polar organic chemical integrative samplers were exposed to surface water for 6 weeks. Alongside the passive samplers an in-situ daphnid test was performed. Subsequent to field exposure, accumulated compounds were extracted from the passive samplers after which a battery of in vivo and in vitro bioassays was exposed to the extracts. The bioassay battery was selected such that it could identify the risks posed by a wide range of chemical pollutants and their transformation products, while simultaneously allowing for targeted identification of groups of compounds that cause specific effects. Bioassay responses were compared to effect-based trigger values to identify potential ecotoxicological risks at the investigated locations. Responses were observed in all bioassays, and trigger values were exceeded in 9 out of the 21 applied assays, allowing for ranking of the investigated locations based on ecotoxicological risks. No relationship between land use and the identification of ecotoxicological risks was observed. Based on the results, considerations regarding future improvements of effect-based monitoring are given. It is concluded that effect-based water quality assessment allowed prioritization of sites based on ecotoxicological risks, identified the presence of hazardous compounds regardless of being listed as priority substances, and meanwhile could prevent costly chemical analysis at sites with low ecotoxicological risks.

Toxicological and ecotoxicological evaluation of the water quality in a large and eutrophic freshwater lake of China

Effect-based methods (EBMs) are recommended as holistic approach for diagnosis and monitoring of water quality; however, the application of EBMs is still scare in China. In the present study, water quality of the freshwater lake Taihu (China) was investigated by EBMs. Different types of water samples were collected from three bays of the lake during 2015, 2016 and 2017. A battery of seven effect-based bioassays, including both specific and non-specific toxicity assays, was used. The bioassay battery was recently suggested based on joint activities of the EU project SOLUTIONS and the NORMAN network on emerging pollutants and is also under discussion for being implemented into monitoring activities in the context of the European Water Framework Directive (WFD). Adverse effects were observed towards the primary producer, primary consumer and fish, indicating the potential ecotoxicity of water in Taihu Lake. Mutagenic and estrogenic effects were found in the Ames fluctuation assay and ERα CALUX (Chemically Activated Luciferase Gene-eXpression) assay, respectively, highlighting the potential risks on human health. Algal growth inhibition and mutagenic effects can be observed during each of the three years. Acute toxicity towards Daphnia magna and estrogen receptor agonistic effects were found in at least one of the samples collected in 2016 and 2017, but not in 2015. The endpoints for fish toxicity in the Danio rerio fish embryo test included both lethal and additionally several sublethal effects (only for samples from 2017) and were not compared between years. Algal growth inhibition, fish embryo toxicity, mutagenic effect and estrogenicity were observed in each of the three bays, while Daphnia acute toxicity was only found in Zhushan Bay. Taking together, this study provides a big picture on the water quality of Taihu Lake. The battery of effect-based tools is promising to be a routine for water quality monitoring in China.

Congo red dye diversely affects organisms of different trophic levels: a comparative study with microalgae, cladocerans, and zebrafish embryos.

Global consumption of synthetic dyes is roughly 7 × 105 tons per year, of which the textile industry expends about two-thirds. Consumption of synthetic dyes produces large volumes of wastewater discharged into aquatic ecosystems. Colored effluents produce toxic effects in the hydrobionts, reduce light penetration, and alter the photosynthetic activity, causing oxygen depletion, among other effects. Some dyes, such as Congo red (CR), are elaborated with benzidine, a known carcinogenic compound. Information regarding dye toxicity in aquatic ecosystems is scarce; therefore, our study was aimed at evaluating the toxicity of CR on a battery of bioassays: the microalga Pseudokirchneriella subcapitata, the cladocerans Daphnia magna and Ceriodaphnia rigaudi, and the zebrafish Danio rerio. P. subcapitata was the most sensitive species to CR (IC50, 3.11mgL-1); in exposed individuals, population growth was inhibited, but photosynthetic pigments and macromolecule concentrations were stimulated. D. magna was tolerant to high dye concentrations, the determined LC50 (322.9mgL-1) is not an environmentally relevant value, but for C. rigaudi, LC50 was significantly lower (62.92mgL-1). In zebrafish embryos, exposure to CR produced yolk sac edema, skeletal deformities, and stopped larvae hatching; lack of heart beating was the only observed lethal effect. CR affected organisms of different trophic levels diversely. Particularly, the effects observed in microalgae confirm the vulnerability of primary producers to dye-polluted wastewaters, because dyes produced toxic effects and interfered with photosynthesis. Different cladoceran species displayed different acute effects; thus, species sensitivity must also be considered when toxicity of dyes is assessed. Inhibition of fish larvae hatching is a significant effect not previously reported that warns about the toxicity of dyes in fish population dynamics. Synthetic azo colorants should be considered as emerging pollutants because they are discharged into the aquatic environment and are not currently included in the environmental regulation of several countries.

Effect-based methods are key. The European Collaborative Project SOLUTIONS recommends integrating effect-based methods for diagnosis and monitoring of water quality

The present monitoring and assessment of the chemical status of water bodies fail to characterize the likelihood that complex mixtures of chemicals affect water quality. The European Collaborative Project SOLUTIONS suggests that this likelihood can be estimated with effect-based methods (EBMs) complemented by chemical screening and/or impact modeling. These methods should be used to identify the causes of impacted water quality and to develop programs of measures to improve water quality. Along this line of reasoning, effect-based methods are recommended for Water Framework Directive (WFD) monitoring to cover the major modes of action in the universe of environmentally relevant chemicals so as to evaluate improvements of water quality upon implementing the measures. To this end, a minimum battery of bioassays has been recommended including short-term toxicity to algae, Daphnia and fish embryos complemented with in vitro and short-term in vivo tests on mode-of-action specific effects as proxies for long-term toxicity. The likelihood of adverse impacts can be established with effect-based trigger values, which differentiate good from poor water quality in close alignment with Environmental Quality Standards for individual chemicals, while taking into account mixture toxicity. The use of EBMs is suggested in the WFD as one avenue to establish the likelihood of adverse effects due to chemical pollution in European water systems. The present paper has been written as one component of a series of policy briefs to support decisions on water quality monitoring and management under the WFD.

Treated acid mine drainage and stream recovery: Downstream impacts on benthic macroinvertebrate communities in relation to multispecies toxicity bioassays

The success and long term effectiveness of extensive and expensive engineering solutions to restore streams impacted by Acid Mine Drainage (AMD) is rarely tested. Concentrations of pollutants were measured in water along a longitudinal gradient from a stretch of the Tweelopie stream, South Africa, that receives pH-treated acid mine drainage (AMD) from an abandoned gold mine. The biotoxic effects of treated AMD were determined through macroinvertebrate biotic indices (SASS5) and a battery of toxicity bioassays. These included the L. sativa, A. cepa, D. magna toxicity and Ames mutagenicity tests, as well as an in vitro human liver cancer cell line HepG2. Even though the Tweelopie stream was moderately to severely degraded by multiple anthropogenic stressors, the impact of the treated AMD was masked by the improvement in the system downstream after mixing with the domestic wastewater effluent receiving stream, and subsequent further dilution as a result of the karst springs downstream. The general improvement of the system downstream was clearly shown by the decrease in the ecotoxicity and mutagenicity in relation to the in-stream macroinvertebrates. PCA multivariate analysis successfully displayed associations between the different environmental variables and the decrease in toxicity and subsequent ecosystem improvement downstream. This study indicated that environmental management of AMD remediation should consider long term assessment strategies, including multiple factors, to promote biological ecosystem recovery.

Cyto/genotoxicological evaluation of hot spots of soil pollution using Allium bioassays in relation to geochemistry

Soil from industrial and landfill sites affected by anthropogenic activity was screened for implicit negative effects in an Allium test-system in relation to geochemistry. The concentrations of 15 elements were compared to the ecotoxicologically-based soil guideline values. Admitted geoindices were used to classify test-soils according to risk/hazard categories. Test-soils were screened for the possible deleterious effects in common onion (Allium cepa L.) by employing a test battery of cytogenetic bioassays (root growth inhibition, mitotic activity, frequency of chromosome aberrations and micronuclei, and cell death rate) complemented with two assays of molecular DNA markers, random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR). Soil from industrial sites was more severely polluted and more cytotoxic for onions compared to soil from landfill sites. However, the cyto/genotoxic outcome of soil exposure in A. cepa was the same for all test-soils; the detrimental effects were observed in onions treated with every test-soil. Thus, test-soils could not be classified as non- and genotoxic, although certain of them had permissible contamination levels. The chromosome aberration frequency and cell death rates were consistent with the intensity of soil contamination, contrary to the micronuclei rate, which was independent of the soil risk/hazard level. Despite a relationship between risk (RI) and total soil contamination (Z) geoindices, both indices correlated with a different Allium cyto/genotoxicity endpoint, although the Z index was preferred over the RI index as being more informative in correlation analysis. Allium bioassays complemented each other by depicting different aspects of exposure to toxic substances, and determination of cyto/genotoxicity in a battery of different bioassays is important in the risk assessment of ecologically dangerous soils, and an application of a test battery is strongly advised.

Effects of sewage sludge application on unfertile tropical soils evaluated by multiple approaches: A field experiment in a commercial Eucalyptus plantation

Sewage sludge (SS) reuse in forest plantation as soil fertilizer/amendment has tremendously increased in recent years. However, SS may have high concentrations of potentially toxic elements (PTE), representing a potential risk for soil and the whole ecosystem. This paper was aimed to assess the toxicity of PTE in unfertile tropical soils amended with SS in a commercial Eucalyptus plantation, with an integrated multiple approaches combining: i) the use of a battery of bioassays (Daphnia magna, Pseudokcrichirella subcapitata, Lactuca sativa, and Allium cepa); and ii) the evaluation of some PTE (Cd, Cr, Cu, Fe, Mn, Ni, Pb, and Zn) and their availability into the pedoenvironment. Differences in total and available PTE between SS doses and time of treatments were evaluated using ANOVA; correlations between PTE and bioassays by a sparse partial robust M-regression (SPRM), while multiple correlations among parameters were performed by principal factor analysis (PFA). Results show that PTE contents in soils tended to increase with SS application doses. However this cannot be assumed as a general rule since in all the investigated treatments the PTE concentrations were consistently below both soil natural background concentrations and quality reference values. Bioassays showed a generalized low eco- and genotoxicity of SS with an increase in toxicity at increasing SS doses but with a clear decreasing trend as time went by. A. cepa was the most sensitive bioassay followed by P. subcapitata > D. magna > L. sativa. Overall, the results indicate that in realistic open field conditions SS risk may be lower than expected due to dynamic decrease in PTE toxicity with time after application. This study has an important implication that open-field trials should be strongly encouraged for evaluating environmental risk of SS application in forestry.

Time-Integrative Passive sampling combined with TOxicity Profiling (TIPTOP): an effect-based strategy for cost-effective chemical water quality assessment

This study aimed at demonstrating that effect-based monitoring with passive sampling followed by toxicity profiling is more protective and cost-effective than the current chemical water quality assessment strategy consisting of compound-by-compound chemical analysis of selected substances in grab samples. Passive samplers were deployed in the Dutch river delta and in WWTP effluents. Their extracts were tested in a battery of bioassays and chemically analyzed to obtain toxicity and chemical profiles, respectively. Chemical concentrations in water were retrieved from publicly available databases. Seven different strategies were used to interpret the chemical and toxicity profiles in terms of ecological risk. They all indicated that the river sampling locations were relatively clean. Chemical-based monitoring resulted for many substances in measurements below detection limit and could only explain <20% of the observed in vitro toxicity. Effect-based monitoring yielded more informative conclusions as it allowed for ranking the sampling sites and for estimating a margin-of-exposure towards chronic effect ranges. Effect-based monitoring was also cheaper and more cost-effective (i.e. yielding more information per euro spent). Based on its identified strengths, weaknesses, opportunities, and threats (SWOT), a future strategy for effect-based monitoring has been proposed.

Design, synthesis, and biological evaluations of phenylpropiolic acid derivatives as novel GPR40 agonists

GPR40, also known as free fatty acid receptor 1 (FFAR1), is a member of G protein-coupled receptors (GPCR) family and has emerged as an attractive target for the treatment of type 2 diabetes mellitus. So far, most of the synthetic GPR40 agonists, including several drug candidates discontinued in clinical trials, were derived from the phenylpropionic acid scaffold. For discovering novel GPR40 agonists with diverse chemical structures, a series of phenylpropiolic acid derivatives were designed, synthesized, and evaluated under a battery of bioassays. Compound 9, the most potent compound in this series, exhibited submicromolar agonist activity and similar agonistic efficacy compared to that of TAK-875. In addition, compound 9 was able to dose-dependently amplify glucose-stimulated insulin secretion (GSIS) in pancreatic β-cell line MIN6, which could be reversed by a selective GPR40 antagonist GW1100. In addition, compound 9 was found to have potent glucose-lowering effects during an oral glucose tolerance test in normal C57BL/6 mice.