Effect-directed investigation and interactive effect of organic toxicants in landfill leachates combining Microtox test with RP-HPLC fractionation and GC/MS analysis

Landfill leachates contain a large amount of unknown harmful compounds derived from domestic and industrial sources. A toxicity effect-directed approach was used to identify biologically active compounds in three landfill leachate samples (S1–S3) by combining the Microtox test with reverse-phase high performance liquid chromatography (RP-HPLC) fractionation and gas chromatography/mass spectrometry (GC/MS) analysis. Organic toxicants were recovered from coarse fractions only in S1 and in S2. Fine fractionation exhibited a somewhat different toxicity pattern in S1 and S2. GC/MS analysis positively identified Bisphenol A (BPA) and 4-t-butylphenol (4-t-BP) in both samples, N-ethyltoluenesulfoneamide (NETSA) was detected only in S1. However, their concentrations were not high enough to be responsible for the observed toxicity in original samples. A synergistic effect among detected organic compounds (BPA, NETSA, and 4-t-BP) was demonstrated. Each compound present at 1/7 of its individual EC₅₀, might lead to undesirable mixture toxicity, which indicated that interactive effects may, to a certain extent, play a role in landfill leachates with complex matrices. The results from further hydrophobicity analysis and estrogen receptor (ER) competitive binding assays of fraction 13 of both samples gave evidence that some possible toxicants that failed to be identified by GC/MS might be endocrine disrupting chemical(s) (EDC) with a log K _ow range of 3.5–3.7 in both samples.     

Chemical composition and toxicity of waste dump leachates using Selenastrum capricornutum Printz (Chlorococcales,Chlorophyta)

Chemical composition and toxicity of leachates from a municipal waste dump in Buenos Aires Province were analyzed. Three sets of samples, obtained in 1996 and 1997, were compared. Levels of nitrogen, phosphorus, and organics (expressed as COD) were distinctly lower in 1997. Such differences were mainly attributed to higher rainfall in that year. The Selenastrum capricornutum assay showed a high sensitivity to heavy metals (Cd=0.049 mg L⁻¹; Cr hexavalent=0.247 mg L⁻¹ and Zn=0.037 mg L⁻¹). Cu, Zn, and Pb concentrations were toxic to algae in leachate samples of the first year, while the EC₅₀ leachates were slightly toxic (86.6 and 99.31%). Most likely, high levels of organics (COD=4640 and 3470 mg L⁻¹) form complex mixtures with metals reducing their toxicity. The EC₅₀ leachate=48.5% in the last year may be explained by high concentrations of Pb and Cu. © 2000 John Wiley & Sons, Inc. Environ Toxicol 15: 76–80, 2000     

Use of Microtox® and Ceriodaphnia bioassays in wastewater fractionation

Collection system and nonchlorinated secondary effluent samples from a large municipal wastewater system were fractionated using a scheme that included filration, EDTA treatment, C₁₈ solid-phase extraction columns, and air stripping. Microtox required less time than Ceriodaphnia dubia bioassay for determining the toxicity of the numerous test samples generated by the fractionation procedure. Its usefulness was limited to collection system samples, however. Secondary effluent samples, which caused significant mortality of C. dubia, were nontoxic to Microtox. Diazinon was tentatively identified as one of the causative toxicants present. Its LC₅₀ to C. dubia (0.5 μg/L) is within the range of concentrations detected (0.1–0.6 μg/L), whereas the EC₅₀ of diazinon to Microtox is much higher (> 18,000 μg/L).     

Cytogenetic effects of leachates from tannery solid waste on the somatic cells of Vicia faba

The contamination of surface- and groundwater by the leaching of solid wastes generated by industrial activities as a result of water runoff and rainfall is a matter of great concern. The leachates from tannery solid waste (TSW), a major environmental pollutant, were examined for their possible genotoxic effects on the somatic cells of Vicia faba. Leachates were prepared from solid wastes procured from leather-tanning industrial sites, and V. faba seedlings were exposed to three test concentrations, 2.5%, 5%, and 10%, through soil and aqueous media for 5 days. The root tips examined for cytogenetic damage revealed that leachate of TSW significantly inhibited the mitotic index and induced significantly frequent chromosomal and mitotic aberrations (CA/MA) in a dose-dependent manner. The chemical analysis of TSW samples revealed that the chief constituents were chromium and nickel, which may cause genetic abnormalities. The frequency of aberrations was found to be higher in the root meristematic cells of Vicia faba exposed through the aqueous medium than those exposed through the soil medium. The results of the present study indicated that contamination of potable water bodies by leachates of TSW may cause genotoxicity. For the biomonitoring of complex mixtures of toxicants with the V. faba bioassay, the use of the aqueous medium seems to be a more promising method than the use of the soil medium.     

Toxicity tests using the kelp <Emphasis Type="Italic">Undaria pinnatifida</Emphasis> for heavy metal risk assessment

The toxicity of six metals (Cd, Cu, Hg, Ni, Pb and Zn) to the early life stages of the brown seaweed Undaria pinnatifida were tested using two endpoints: spore germination and germ tube elongation. The optimal test conditions determined for photon irradiance, pH, salinity and temperature were darkness, pH 8, 35‰ and 15°C, respectively. The EC₅₀ values could be determined only for Hg and Cu and were 27 μg · L^-1 and 186 μg · L^-1, respectively, for germination and 16 μg · L^-1 and 45 μg · L^-1, respectively, for germ tube elongation. The variability of our U. pinnatifida test in response to the metal toxicants, as estimated from the coefficient of variation of the mean EC₅₀ values, was less than 28%. When tested against two different wastewater samples (PCB manufacturer and TV & communication equipment manufacturer), U. pinnatifida exhibited an EC₅₀ of 66.09-80.73% for germination and 24.98-62.62% for germ tube elongation with the CV (Coefficient of variation) range for both germination and germ tube elongation between 1.24 and 13.18%. The novel aspect of the present method is that the testing procedure requires no lighting or photoperiodic control. Additionally, the response of Undaria to the toxicity of Hg and Cu is very similar to that reported for other very sensitive macroalgal species. Because of its ecological and economic importance, the evaluation of toxicity using early stages of U. pinnatifida will provide useful scientific information, and we propose that this represents a valid bioassay within the battery of aquatic bioassays for the assessment of anthropogenic impacts on coastal ecosystems and commercial cultivation areas in nearshore environments.     

Adapting an enzymatic toxicity test to allow comparative evaluation of natural freshwater biofilms’ tolerance to metals

A simple, low-cost and non-radioactive short-term toxicity test was developed to study the effects of urban metals on natural freshwater periphytic communities. β-glucosidase activity of natural freshwater biofilms collected in situ was chosen as an endpoint. Metals (Cd, Cu, Ni, Pb, and Zn) successfully inhibited bacterial enzymatic activity after a 1-h exposure enabling the calculation of EC₅₀. The EC₅₀ value of a biofilm sample varied with the Total Suspended Solids concentration (TSS) of the biofilm suspension, showing that EC₅₀ values (expressed as total added metal concentrations) are not representative of the bioavailable metal concentration during the toxicity test. For Cu, Cd, Ni, Zn and Pb, the EC₅₀ values increased linearly with the TSS concentration leading us to define a normalized EC₅₀: the value of the EC₅₀ divided by the corresponding TSS concentration. Normalized EC₅₀ proved to be a robust, reliable way to assess metal tolerance of a biofilm for Cd, Cu, Ni, Zn and Pb. Normalized EC₅₀ obtained, expressed as kg_metal/g_TSS, varied between 0.2 to 7.6 for Cu, 1 to 8 for Cd, 1.8 to 92.3 for Ni, 1.8 to 76.6 for Zn and 25 to 189 for Pb.     

An assessment of the potential of the microbial assay for risk assessment (MARA) for ecotoxicological testing

Rapid microscale toxicity tests make it possible to screen large numbers of compounds and greatly simplify toxicity identification evaluation and other effect directed chemical analyses of effluents or environmental samples. Tests using Vibrio fischeri (such as Microtox?) detect toxicants that cause non-specific narcosis, but are insensitive to other important classes of contaminants. The microbial assay for risk assessment (MARA) is a 24 h multi-species test that seeks to address this problem by using a battery of ten bacteria and a fungus. But there has been little independent evaluation of this test, and there is no published information on its sensitivity to pesticides. Here, we assess the performance of MARA using a range of toxicants including reference chemicals, fungicides and environmental samples. Mean MARA microbial toxic concentrations and IC₂₀s (20% Inhibitory concentrations) indicate the toxicant concentrations affecting the more sensitive micro-organisms, while the mean IC₅₀ (50% Inhibitory concentration) was found to be the concentration that was toxic to most MARA species. For the two fungicides tested, the yeast (Pichia anomalia) was the most sensitive of the ten MARA species, and was more sensitive than the nine other yeasts tested. The test may be particularly valuable for work with fungicides. Mean MARA IC₅₀s were comparable to values for nine other yeast species and the lowest individual IC₅₀s for each toxicant were comparable to reported IC₅₀s for Daphnia magna, Selenastrum capricornutum and Microtox? bioassays. MARA organisms exhibited more variable sensitivities, with the most sensitive organism being different for different samples, enhancing the likelihood of toxicity detection and giving a toxicity “fingerprint” that may help identify toxicants. The test, therefore, has great potential and would be valuable for ecotoxicological testing of pollutants.     

Species-specific sensitivity to selenium-induced impairment of cortisol secretion in adrenocortical cells of rainbow trout (Oncorhynchus mykiss) and brook trout (Salvelinus fontinalis)

Species differences in physiological and biochemical attributes exist even among closely related species and may underlie species-specific sensitivity to toxicants. Rainbow trout (RT) are more sensitive than brook trout (BT) to the teratogenic effects of selenium (Se), but it is not known whether all tissues exhibit this pattern of vulnerability. In this study, primary cultures of RT and BT adrenocortical cells were exposed to selenite (Na₂SO₃) and selenomethionine (Se-Met) to compare cell viability and ACTH-stimulated cortisol secretion in the two fish species. Cortisol, the primary stress hormone in fish, facilitates maintenance of homeostasis when fish are exposed to stressors, including toxicants. Cell viability was not affected by Se, but selenite impaired cortisol secretion, while Se-Met did not (RT and BT EC₅₀ > 2000 mg/L). RT cells were more sensitive (EC₅₀ = 8.7 mg/L) to selenite than BT cells (EC₅₀ = 90.4 mg/L). To identify the targets where Se disrupts cortisol synthesis, selenite-impaired RT and BT cells were stimulated with ACTH, dbcAMP, OH-cholesterol, and pregnenolone. Selenite acted at different steps in the cortisol biosynthesis pathway in RT and BT cells, confirming a species-specific toxicity mechanism. To test the hypothesis that oxidative stress mediates Se-induced toxicity, selenite-impaired RT cells were exposed to NAC, BSO and antioxidants (DETCA, ATA, Vit A, and Vit E). Inhibition of SOD by DETCA enhanced selenite-induced cortisol impairment, indicating that oxidative stress plays a role in Se toxicity; however, modifying GSH content of the cells did not have an effect. The results of this study, with two closely related salmonids, provided additional evidence for species-specific differences in sensitivity to Se which should be considered when setting thresholds and water quality guidelines.     

Performance of a miniaturized algal bioassay in phytotoxicity screening

A miniaturized and low-cost algal growth-inhibition assay, with Pseudokirchneriella subcapitata, based on the standard ISO 8692 and using 96-well microplates, was tested and optimized in this work, to be used as a useful tool for pollutant phytotoxicity screening. For validation, the performance of the microplate algal growth-inhibition assay was first compared with the standard flask assay for the toxicity testing of five reference toxicants (copper(II) sulfate, zinc sulfate, potassium permanganate, potassium dichromate and 3,5-Dichlorophenol) and six wastewater samples. Statistical evaluation of EC₅₀ results from both methods demonstrated a good agreement between microplate and flask assays either in testing chemicals (r ² = 0.975, p < 0.0017) or environmental samples toxicity (r ² = 0.984, p < 0.0001). In addition, the performance of this algal microplate bioassay was also evaluated in comparison with Lemna test, ISO 20079, for phytotoxicity assessment of 27 wastewater samples from industries and treatment plants. The results showed that the algal test was more sensitive for most of the samples, but a significant agreement between both tests was observed (r ² = 0.644, p < 0.0001). In conclusion, this miniaturized test can be a good tool to include in a battery of tests for phytotoxicity screening of a wide range of chemicals and environmental samples, with the advantage of requiring low sample volumes for the test, allowing large numbers of samples to be tested, and generating low volumes of waste.     

Pharmacological studies confirm neurotoxic metabolite(s) produced by the bloom‐forming Cylindrospermopsis raciborskii in Hungary

A rapid cyanobacterial bloom of Cylindrospermopsis raciborskii (3.2 × 10⁴ filaments/mL) was detected early November, 2012, in the Fancsika pond (East Hungary). The strong discoloration of water was accompanied by a substantial fish mortality (even dead cats were seen on the site), raising the possibility of some toxic metabolites in the water produced by the bloom‐forming cyanobacteria (C. raciborskii). The potential neuronal targets of the toxic substances in the bloom sample were studied on identified neurons (RPas) in the central nervous system of Helix pomatia. The effects of the crude aqueous extracts of the Fancsika bloom sample (FBS) and the laboratory isolate of C. raciborskii from the pond (FLI) were compared with reference samples: C. raciborskii ACT 9505 (isolated in 1995 from Lake Balaton, Hungary), the cylindrospermopsin producer AQS, and the neurotoxin (anatoxin‐a, homoanatoxin‐a) producer Oscillatoria sp. (PCC 6506) strains. Electrophysiological tests showed that both FBS and FLI samples as well the ACT 9505 extracts modulate the acetylcholine receptors (AChRs) of the neurons, evoking ACh agonist effects, then inhibiting the ACh‐evoked neuronal responses. Dose–response data suggested about the same range of toxicity of FBS and FLI samples (EC₅₀ = 0.397 mg/mL and 0.917 mg/mL, respectively) and ACT 9505 extracts (EC₅₀ = 0.734 mg/mL). The extract of the neurotoxin‐producing PCC 6506 strain, however, proved to be the strongest inhibitor of the ACh responses on the same neurons (EC₅₀ = 0.073 mg/mL). The presented results demonstrated an anatoxin‐a‐like cholinergic inhibitory effects of cyanobacterial extracts (both the environmental FBS sample, and the laboratory isolate, FLI) by some (yet unidentified) toxic components in the matrix of secondary metabolites. Previous pharmacological studies of cyanobacterial samples collected in other locations (Balaton, West Hungary) resulted in similar conclusions; therefore, we cannot exclude that this chemotype of C. raciborskii which produce anatoxin‐a like neuroactive substances is more widely distributed in this region. © 2013 Wiley Periodicals, Inc. Environ Toxicol 30: 501–512, 2015.     

A simple microplate algal assay technique for aquatic toxicity assessment

A simple, miniaturized algal assay procedure using microplates has been developed to assess aquatic toxicity with the green alga, Selenastrum capricornutum. Cell count-generated EC₅₀ data comparisons with the classic assay using flasks have indicated good agreement between the two methods following toxic assessment of various wastewater samples and metal solutions. Parametric comparisons (ATP us cell counts) with the microplate method show equally good correlation. The technique is highly versatile in conducting basic algal bioassays for varying times (4-hour, 4-day, 8-day EC₅₀'s) and with differing parameters, depending on set objectives. Other interesting features of the microplate technique include handling rapidity, economy, space-saving convenience, and automation potential.     

Toxicity analysis of leachates from hazardous wastes via microtox and daphnia magna

One of the most common ways of surface and ground water contamination from hazardous wastes is through its leachates. Toxicity is a meaningful parameter allowing an integrated evaluation of the potential danger of leachates. Two bioassays were considered for measuring toxicity: Microtox and Daphnia magna. The toxicity was measured on leachates obtained, using different leaching procedures, from wastes of a pesticide manufacturing industry and sludge from electroplating wastewater. Twenty-two tests were carried out, measuring IC50 for D. magna and EC50 for Microtox. Both bioassays were compared using three toxicity criteria. The first criterium is classification on toxic/non-toxic at 25 and 50% leachate concentration. The second criterium is classification on percent ranks, and the third on log ranks. Considering these criteria, the agreement between both bioassays is within a 75–85%. It is shown that both, Microtox and D. magna assays could be used as toxicity indicators for the wastes considered. The sensitivity of the bioassays is different depending on leachate composition. In all samples, the leachate concentration of chemicals was measured.     

Influence of salinity on metal toxicity to Ulva pertusa

The present study was undertaken to establish the effect of salinity on the toxicity of cadmium (Cd), copper (Cu), lead (Pb) and zinc (Zn) to Ulva pertusa, with inhibition of spore release used as the endpoint. The optimal salinity for maximal spore release for U. pertusa was found to be between 20 and 40 psu. Comparisons between toxicity of metals, as measured by EC₅₀, was shown to be in the descending order of Cu>Cd>Pb=Zn, which is similar to the toxicity of metals to algae, in general. When salinity was decreased from 30 to 20 psu, the EC₅₀ values for Cd toxicity to the inhibition of spore release in U. pertusa decreased from 261 to 103 g·L^?1, whereas increased salinity from 30 to 40 psu increased the EC₅₀ from 261 to 801 g·L^?1. Similarly, EC₅₀ values for Cu toxicity were 52 g·L^?1 at 20 psu, 99 g·L^?1 at 30 psu, and 225 g·L^?1 at 40 psu, and for Zn toxicity were 720 g·L^?1, 1,074 g·L^?1 and 1,520 g·L^?1, at 20, 30 and 40 psu, respectively. In contrast, no salinity dependent change in EC₅₀ values was apparent for Pb, with no significant differences in EC₅₀ values at under the three different salinity regimes. In general, lower salinity (20 psu) induced a significant decrease in percent spore release of U. pertusa as estimated by a decrease in EC₅₀ values, while higher salinity (40 psu) reduced the toxicity of metals as shown by an increase EC₅₀ values. These findings enable one to predict that any additional increase in pollution status would result in a pronounced reduction in the distribution of U. pertusa in brackish and estuarine waters.     

Acute aquatic toxicity of tire and road wear particles to alga,daphnid, and fish

Previous studies have indicated that tire tread particles are toxic to aquatic species, but few studies have evaluated the toxicity of such particles using sediment, the likely reservoir of tire wear particles in the environment. In this study, the acute toxicity of tire and road wear particles (TRWP) was assessed in Pseudokirchneriella subcapita, Daphnia magna, and Pimephales promelas using a sediment elutriate (100, 500, 1000 or 10000 mg/l TRWP). Under standard test temperature conditions, no concentration response was observed and EC/LC₅₀ values were greater than 10,000 mg/l. Additional tests using D. magna were performed both with and without sediment in elutriates collected under heated conditions designed to promote the release of chemicals from the rubber matrix to understand what environmental factors may influence the toxicity of TRWP. Toxicity was only observed for elutriates generated from TRWP leached under high-temperature conditions and the lowest EC/LC₅₀ value was 5,000 mg/l. In an effort to identify potential toxic chemical constituent(s) in the heated leachates, toxicity identification evaluation (TIE) studies and chemical analysis of the leachate were conducted. The TIE coupled with chemical analysis (liquid chromatography/mass spectrometry/mass spectrometry [LC/MS/MS] and inductively coupled plasma/mass spectrometry [ICP/MS]) of the leachate identified zinc and aniline as candidate toxicants. However, based on the high EC/LC₅₀ values and the limited conditions under which toxicity was observed, TRWP should be considered a low risk to aquatic ecosystems under acute exposure scenarios.     

The effect of ionic solutes on EC50 values measured using the microtox test

The Microtox test uses the marine luminescent bacterium, Photobacterium phosphoreum. To provide the organism with osmotic protection, sodium chloride (2.0%) is usually added to freshwater samples before analysis. This procedure is shown to cause a dramatic lowering of measured toxicity where ionic compounds are the major toxicants present. The reason for this is the consequent variation in ionic strength upon addition of NaCl rather than formation of less-toxic chloro complexes. Data are given for the toxicants Zn, Cd, Cu, NH₄⁺, and phenol. It was found that measurements on aqueous samples could be made without adjustment provided that the salinity was in the range of 1.0–7.0%. Freshwaters can also be analyzed without adjustment of ionic strength by adding sucrose (20.4%) to provide the necessary osmotic protection. It is therefore recommended that the ionic strength of aqueous samples should not be altered as part of the test and that receiving waters should be used as the diluent wherever possible.     

What are the differences between aerobic and anaerobic toxic effects of sulfonamides on Escherichia coli?

Bacteria in the environment face the threat of antibiotics. However, most studies investigating the toxicity and toxicity mechanisms of antibiotics have been conducted on microorganisms in aerobic conditions, while studies examining the anaerobic toxicity and toxicity mechanisms of antibiotics are still limited. In this study, we determined the aerobic and anaerobic toxicities of sulfonamides (SAs) on Escherichia coli. Next, a comparison of the aerobic and anaerobic toxicities indicated that the SAs could be divided into three groups: Group I: log(1/EC_50-anaerobic) > log(1/EC_50-aerobic) (EC_50-anaerobic/EC_50-aerobic, the median effective concentration under anaerobic/aerobic conditions), Group II: log(1/EC_50-anaerobic) ≈ log(1/EC_50-aerobic), and Group III: log(1/EC_50-anaerobic) < log(1/EC_50-aerobic). Furthermore, this division was not based on the reactive oxygen species (ROS) level or the interaction energy (E_binding) value, which represents the affinity between SAs and dihydropteroate synthase (dhps) but rather on the total binding energy. Furthermore, SAs with greatly similar structures were categorized into different groups. This deep insight into the difference between aerobic and anaerobic toxicities will benefit environmental science, and the results of this study will serve as a reference for the risk assessment of chemicals in the environment.     

Evaluation of consistency for multiple experiments of a single combination in the time-dependence mixture toxicity assay

Test-to-test consistency was evaluated for a single binary combination of organic chemicals using an assay that examined toxicity over multiple exposure times. Six experiments were conducted. The toxicities of 3-chloro-2-butanone (3C2B), methyl crotonate (MC) and the mixture of both (MX) were evaluated in each experiment at 15, 30 and 45?min of exposure using the Microtox^® system. Concentration-response (x/y) curves were generated via the five-parameter logistic minus one-parameter (5PL???1P) curve-fitting function and were used to develop predicted x/y curves for the dose-addition (DA) and independence (I) models of combined effect. A variety of toxicity (e.g. effective concentration, EC₅₀) and time-dependent toxicity (TDT) endpoints, 5PL???1P parameters and various combined-effects metrics (e.g. MX/DA) were calculated. Test-to-test consistency was evaluated via the coefficient of variation (CV) or, for TDT, the standard deviation of mean values. In the study, CVs obtained for single-chemical and mixture toxicity endpoints (EC₂₅, EC₅₀ and EC₇₅) at each exposure time were?<20, as were those for the predicted DA and I curves. For the MX/DA metric, mixture toxicity was consistent with that predicted for DA at each exposure time in each experiment with CVs?<6, despite some substantial variation in TDT for MC-alone at the EC₂₅ and for the 30–45?min time-interval. There was a lower variation in TDT for 3C2B and MX. Mean and CV values for 5PL???1P-derived slope and asymmetry parameters were also assessed to provide bases for comparisons in future reports.     

Pollution-induced community tolerance of freshwater biofilms: measuring heterotrophic tolerance to Pb using an enzymatic toxicity test

This study aims at investigating the impacts of Pb on freshwater biofilms with a pollution-induced community tolerance (PICT) approach using a recently developed short-term toxicity test based on β-glucosidase activity to measure biofilms’ tolerance to Pb. We first investigated more closely the influence of the total suspended solids (TSS) concentrations of biofilm suspensions used for short-term toxicity tests performed to assess Pb tolerance. The Pb EC₅₀ values of four dilutions of the same biofilm suspension increased with their TSS concentrations. TSS-normalization allowed to obtain a unique measure of Pb tolerance, thus confirming that TSS-normalization of EC₅₀ values is a good means to estimate biofilm tolerance to Pb. The experiment was repeated with three different biofilm samples collected at different sites and dates. Second, biofilms were exposed to Pb (0, 1, 10 and 100?μg/L) for 3?weeks in microcosms to assess the impacts of Pb exposure on the communities. An increase in Pb tolerance was observed for the biofilm exposed to 100?μg/L. Automated Ribosomal Intergenic Spacer Analysis revealed modifications of bacterial and eukaryotic community structure with Pb exposure. Moreover, exposure to 100?μg/L Pb also led to an increase in Zn tolerance but not Cu tolerance. This study shows that tolerance acquisition to Pb can be detected after exposure to environmental concentrations of Pb using a PICT methodology and normalized EC₅₀ values as measures of Pb tolerance.     

Cell death and inhibition of population growth of marine unicellular algae by pesticides

The marine diatom, Skeletonema costatum, was exposed to the pesticides hexachlorocyclopentadiene, EPN, chlorpyrifos, carbophenothion, and Atrazine and examined for death of cells with Evans blue, a mortal stain. All pesticides caused death of cells, but significant mortality occurred only at concentrations greater than the EC₅₀ calculated from population growth studies. The insecticide, Amdro^®, did not kill the marine algae S. costatum, Thalassiosira pseudonana, Isochrysis galbana, Chlorella sp., or Dunaliella tertiolecta. However, Amdro was very inhibitory to algal population growth at low concentrations, i.e. EC₅₀ values at 48 h were between 0.14 ppb for T. pseudonana and 10.3 ppb for D. tertiolecta. EC₅₀ values were lowest after 48 h of exposure in tests conducted for 96 h. By 96 h after exposure, maximum growth rates of treated cultures were approximately those of control cultures. It is recommended that, because of problems associated with the fate of toxicants, algal laboratory toxicity tests be conducted for 48 to 72 h instead of the usual 96 h or longer.     

Acute toxicity of chlorinated organic chemicals to the embryos and larvae of the marine worm Platynereis dumerilii (Polychaeta: Nereidae)

The acute toxicity of a range of organic reference chemicals has been evaluated using the embryo-larval stages of the marine polychaete Platynereis dumerilii (Audouin and Milne-Edwards). After exposure under static conditions at 20±1°C, EC₅₀ values were calculated based upon reductions in normal embryo development. After exposure of 7 day old larvae under the same conditions, LC₅₀ values were calculated based on larval survival. The embryo development 48 h EC₅₀ and larval 48 h LC₅₀ values were, respectively, 6.39 and 8.94 mg L⁻¹ for 3,4-dichloroaniline, 122 and 104 mg L⁻¹ for phenol, 23.6 and 14.4 mg L⁻¹ for 4-chlorophenol, 4.24 and 3.20 mg L⁻¹ for 3,5-dichlorophenol, 2.55 and 4.24 mg L⁻¹ for 2,4,5-trichlorophenol, and 0.52 and 1.33 mg L⁻¹ for sodium pentachlorophenate (all based on nominal values). Comparison of these data with other published values suggest that, for these reference toxicants, the embryo–larval stages of P. dumerilii are of similar sensitivity to the early life stages of several other marine invertebrate species. These observations add to the refinement of a suite of bioassay techniques using P. dumerilii which, linked to previous studies, can be used to evaluate the developmental toxicity, cytotoxicity, and genotoxicity of marine contaminants. © 1998 John Wiley & Sons, Inc. Environ Toxicol Water Qual 13: 149–155, 1998