Single-species Laboratory Tests Research Articles

Evaluation of short-term copper toxicity in a co-culture of Synechococcus sp., Chaetoceros gracilis and Pleurochrisys cf. roscoffensis exposed to changes in temperature and salinity levels

Metals such as copper (Cu) enter marine environments from natural and anthropogenic sources, causing changes in the biodiversity of marine microalgae and cyanobacteria. Cu plays a dual role as either a micronutrient or toxicant depending on the environmental concentration. Many studies have summarized the potential of Cu to become more toxic to microalgae under environmental stress (for instance climate change).Most of the data available on Cu toxicity concerning microalgae and cyanobacteria have been produced using single-species laboratory tests, and there is still a significant gap in the information concerning the behavior of a group of algae exposed to environmental stressors. Thus, the objective of this study was to evaluate the toxicity of Cu at two concentrations (C1 = 2 μg L−1 and C2 = 5 μg L−1) in multispecies bioassays using three phytoplankton species (one cyanobacteria, Synechococcus sp., and two microalgae, Chaetoceros gracilis and Pleurochrisys cf. roscoffensis). Combinations of two temperatures (20 and 23 °C) and two salinities (33 and 36 PSU), were applied in a 96 h study using flow cytometry analysis (FCM). Algal growth and reactive oxygen species (ROS) production by 2′7′-dichlorofluorescein (DCFH) were monitored by FCM. The results indicated that Synechococcus sp. was more sensitive than C. gracilis and P. roscoffensis to Cu stress at a temperature 23 °C and salinity of 36 PSU under both concentrations of Cu. Chlorophyll a fluorescence showed a significant decrease (p < 0.05) in Synechococcus sp. under 5 μg L−1 of Cu in the combined treatment of 20 °C and 33 PSU; however, there was a significant increase in P. roscoffensis in all combinations at C2 = 5 μg L−1 compared to the control with no Cu, indicating a potential hormetic response to Cu for P. roscoffensis. ROS levels were triggered in a combination of 23 °C and 33 PSU and 5 μg L−1 of Cu, which was higher than all the other combinations studied. Our study resulted in data concerning the potential impacts caused by possible future climate change scenarios in aquatic habitats chronically exposed to metals.

Do magnesium and chloride ameliorate high sodium bicarbonate concentrations? A comparison between laboratory and mesocosm toxicity experiments

Increasing salinity is a concern for biodiversity in many freshwater ecosystems globally. Single species laboratory toxicity tests show major differences in freshwater organism survival depending on the specific ions that comprise salinity types and/or their ion ratios. Toxicity has been shown to be reduced by altering ionic composition, despite increasing (total) salinity. For insistence, single species tests show the toxicity of sodium bicarbonate (NaHCO3, which commonly is a large proportion of the salts from coalbeds) to freshwater invertebrates is reduced by adding magnesium (Mg2+) or chloride (Cl−). However, it is uncertain whether reductions in mortality observed in single-species laboratory tests predict effects within populations, communities and to ecosystem processes in more complex multi-species systems both natural and semi-natural. Here we report the results of an outdoor multi-species mesocosm experiment to determine if the effects of NaHCO3 are reduced by increasing the concentrations of Mg2+ or Cl− on: a) stream macroinvertebrate populations and communities; b) benthic chlorophyll-a and; c) the ecosystem process of leaf litter decomposition. We found a large effect of a high NaHCO3 concentration (≈4.45 mS/cm) with reduced abundances of multiple taxa, reduced emergence of adult insects and reduced species richness, altered community structure and increased leaf litter breakdown rates but no effect on benthic chlorophyll-a. However, despite predictions based on laboratory findings, we found no evidence that the addition of either Mg2+ or Cl− altered the effect of NaHCO3. In semi-natural environments such as mesocosms, and natural environments, organisms are subject to varying temperature and habitat factors, while also interacting with other species and trophic levels (e.g. predation, competition, facilitation), which are absent in single species laboratory tests. Thus, it should not be assumed single-species tests are good predictors of the effects of changing ionic compositions on stream biota in more natural environments.

Tiered Approach for the Evaluation of Environmental Impacts of Triclosan on Aquatic Ecosystems

The synthetic broad-spectrum antibacterial agent triclosan is one of the most commonly encountered emerging micro-pollutant in the aquatic environment due to the extensive use since 1968 mainly in cosmetics and household cleaning products and the partial elimination from wastewater. Its low water solubility, high sorption coefficient to organic matter, accumulation potential in fatty tissues and its low acute toxicity determined by conventional ecotoxicological tests suggest that its risk is more related to chronic effects requiring risk assessment based on more sensitive ecotoxicological methods. In this paper the short- and midterm ecotoxicological effects of triclosan were investigated using various test systems taking into account ecological complexity and environmental relevance. Acute single-species, simplified microcosm experiments and complex multi-species microcosm experiments were conducted with the determination of sublethal physiological and behavioral endpoints including the Daphnia magna heartbeat rate and feeding activity, the Heterocypris incongruens movement parameters and the Lemna minor chlorophyll content and root length. All physiological and behavioral endpoints indicated sensitively the adverse effect of triclosan in the concentration range of 4–25.6 µg/L. In some cases, responses of selected organisms in single-species laboratory tests did not correspond to those of the higher levels of test systems. Daphnia sensitivity increased with the level of the test system for all chosen endpoints except the heartbeat rate. Considering the varying ecological complexity of the assembled test systems, according to our results the exposure time and the different combinations of exposure routes were the most decisive parameters in terms of triclosan ecotoxicity and endpoint sensitivity.

Higher than … or lower than ….? Evidence for the validity of the extrapolation of laboratory toxicity test results to predict the effects of chemicals and ionising radiation in the field

Single species laboratory tests and associated species sensitivity distributions (SSDs) that utilise the resulting data can make a key contribution to efforts to prospective hazard assessments for pesticides, biocides, metals and ionising radiation for research and regulatory risk assessment. An assumption that underlies the single species based toxicity testing approach when combined in SSD models is that the assessments of sensitivities to chemical and ionising radiation measured across a range of species in the laboratory can inform on the likely effects on communities present in the field. Potential issues with the validity of this assumption were already recognised by Van Straalen and Denneman (1989) in their landmark paper on the SSD methodology. In this work, they identified eight major factors that could potentially compromise the extrapolation of laboratory toxicity data to the field. Factors covered a range of issues related to differences in chemistry (e.g. bioavailability, mixtures); environmental conditions (optimal, variable), ecological (compensatory, time-scale) and population genetic structure (adaptation, meta-population dynamics). This paper outlines the evidence pertaining to the influence of these different factors on toxicity in the laboratory as compared to the field focussing especially on terrestrial ecosystems. Through radiological and ecotoxicological research, evidence of the influence of each factor on the translation of observed toxicity from the laboratory to field is available in all cases. The importance of some factors, such as differences in chemical bioavailability between laboratory tests and the field and the ubiquity of exposure to mixtures is clearly established and has some relevance to radiological protection. However, other factors such as the differences in test conditions (optimal vs sub-optimal) and the development of tolerance may be relevant on a case by case basis. When SSDs generated from laboratory tests have been used to predict chemical and ionising radiation effects in the field, results have indicated that they may often seem to under-predict impacts, although this may also be due to other factors such as the effects of other non-chemical stressors also affecting communities at polluted sites. A better understanding of the main factors affecting this extrapolation can help to reduce uncertainty during risk assessment.

Assessment of toxicity of heavy metal-contaminated soils toward Collembola in the paddy fields supported by laboratory tests.

Effects on soil Collembola of Cu, Zn, Pb, and Cd pollution from Cu smelters over 40years were investigated in paddy fields from an area of Eastern China. We compared the field effects to those observed in single-species laboratory tests employing the hemiedaphic collembolan Folsomia candida and the epedaphic Sinella curviseta obtained from laboratory cultures and exposed to field-collected polluted soil. The results indicated that different collembolan species responded differently to the pollution in the fields and could be divided into sensitive, indifferent, and tolerant types accordingly. The abundance of sensitive species decreased as the pollution increased, but this was not the same for indifferent and tolerant species. The dominant species changed from sensitive to tolerant species as the pollution increased. The reproduction of F. candida and S. curviseta was most sensitive to the contaminated soil compared to growth and survival; the sensitivity of the two species was similar. The growth was more sensitive than the survival for F. candida but not for S. curviseta. The growth and survival of F. candida were much more sensitive than those of S. curviseta. Sensitivity of field populations of F. candida (EC10 31 [15-46]) and hemiedaphic species Folsomia quadrioculata (EC10 52 [0.7-102]) were comparable with sensitivity of the reproduction of F. candida in the single-species tests (EC10 21 [14-27]), suggesting that single-species test based on laboratory cultures and field soil could be used to link laboratory and field data and then reflect the field situation. S. curviseta could be used as an epedaphic species in single-species tests and F. quadrioculata as an indicator species for assessment of field effect.

Long-Term Recovery of Benthic Communities in Sediments Amended with Activated Carbon

Using activated carbon (AC) for sediment remediation may have negative effects on benthic communities. To date, most AC effect studies were short-term and limited to single species laboratory tests. Here, we studied the effects of AC on the recolonization of benthic communities. Sediment from an unpolluted site was amended with increasing levels of AC, placed in trays and randomly embedded in the original site, which acted as a donor system for recolonization of benthic species. After 3 and 15 months, the trays were retrieved and benthic organisms identified. A positive trend with AC was detected for species abundance after 3 months, whereas after 15 months a negative trend with AC was detected for Lumbriculidae and Pisidiidae. On the community level, statistical analyses showed a considerable recovery in terms of species diversity and abundance in 3 months and full recovery of the community after 15 months. This was explained from migration of individuals from the donor system, followed by further migration and reproduction of the species in the next year. AC treatments explained 3% of the variance in the community data. This work suggests that AC community effects are mild as long as AC levels are not too high (1-4%).

How to test nontarget effects of veterinary pharmaceutical residues in livestock dung in the field

To register veterinary medicinal products (VMPs) as parasiticides on pastured animals, legislation in the European Union requires an environmental risk assessment to test the potential nontarget effects of fecal residues on dung-dwelling organisms. Products with adverse effects in single-species laboratory tests require further, higher-tier testing to assess the extent of these effects on entire communities of dung-dwelling organisms under more realistic field or semifield conditions. Currently, there are no documents specifically written to assist researchers in conducting higher-tier tests or to assist regulators in interpreting the results of such tests in an appropriate context. Here we provide such a document, written by members of the SETAC Advisory Group DOTTS (Dung Organism Toxicity Testing Standardization) with research experience on dung fauna in central and southern Europe, Canada, Australia, and South Africa. This document briefly reviews the organisms that make up the dung community and their role in dung degradation, identifies key considerations in the design and interpretation of experimental studies, and makes recommendations on how to proceed.

Conducting model ecosystem studies in tropical climate zones: Lessons learned from Thailand and way forward

Little research has been done so far into the environmental fate and side effects of pesticides in the tropics. In addition, those studies conducted in tropical regions have focused almost exclusively on single species laboratory tests. Hence, fate and effects of pesticides on higher-tier levels have barely been studied under tropical conditions. To address this lack of knowledge, four outdoor aquatic model ecosystem experiments using two different test systems were conducted in Thailand evaluating the insecticide chlorpyrifos, the herbicide linuron and the fungicide carbendazim. Results of these experiments and comparisons of recorded fate and effects with temperate studies have been published previously. The present paper discusses the pros and cons of the methodologies applied and provides indications for i) possible improvements; ii) important aspects that should be considered when performing model ecosystem experiments in the tropics; iii) future research.

Effects of boric acid on various microbes, plants, and soil invertebrates

Purpose Terrestrial ecotoxicology includes the investigation of the impact of chemicals on soils and soil-inhabiting organisms, whereby ecotoxicological effects are mainly assessed using single-species laboratory tests. As part of laboratory testing, reference substances, with known toxicity, are regularly used to confirm the sensitivity of the cultivated test organisms over time. Furthermore, reference substances are also applied in ring test evaluations for the validation of new test methods. The present work evaluates boric acid for its suitability as a potential reference substance for tests with higher plants and soil organisms (e.g., invertebrates and microbial communities).

The species sensitivity distribution approach compared to a microcosm study: A case study with the fungicide fluazinam

We assessed the sensitivity of freshwater organisms (invertebrates and algae) to the fungicide Shirlan ® (active ingredient fluazinam) in single-species laboratory tests and in microcosms. Species sensitivity distribution (SSD) curves were constructed by means of acute toxicity data for 14 invertebrate species, since algae were much less sensitive. The EC 10-based SSD gave a median HC 5 value of 0.6 μg L −1 and a 90% confidence interval of 0.1–1.9 μg L −1. The EC 50-based SSD gave a median HC 5 value of 3.9 μg L −1 (90% confidence interval: 0.9–9.9 μg L −1). The microcosms were treated four times with Shirlan ® (concentration range: 0.4–250 μg L −1). Responses of the microcosm communities were followed. The 2 μg L −1 treatment was the no-observed-effect concentration (NOEC microcosm). The 10 μg L −1 treatment resulted in short-term effects on a few zooplankton taxa. Clear effects were observed at 50 and 250 μg L −1. The responses in the microcosms were in line with the toxicity data for the tested lab species. The median EC 10-based HC 5 and the lower limit EC 50-based HC 5 were lower, and the median EC 50-based HC 5 was slightly higher than the NOEC microcosm. This is consistent with other studies that compared SSDs with responses in model ecosystems that received repeated applications of pesticides.

Comparative aquatic toxicity of the pyrethroid insecticide lambda-cyhalothrin and its resolved isomer gamma-cyhalothrin

In this review we compare the sensitivity of a range of aquatic invertebrate and fish species to gamma-cyhalothrin (GCH), the insecticidally active enantiomer of the synthetic pyrethroid lambda-cyhalothrin (LCH), in single-species laboratory tests and outdoor multi-species ecosystem tests. Species sensitivity distribution curves for GCH gave median HC(5) values of 0.47 ng/L for invertebrates, and 23.7 ng/L for fish, while curves for LCH gave median HC(5) values of 1.05 ng/L and 40.9 ng/L for invertebrates and fish, respectively. A model ecosystem test with GCH gave a community-level no observed effect concentration (NOEC(community)) of 5 ng/L, while model ecosystem tests with LCH gave a NOEC(community) of 10 ng/L. These comparisons between GCH and LCH indicate that the single active enantiomer causes effects at approximately one-half the concentration at which the racemate causes similar effects.

Effects of the pyrethroid insecticide gamma-cyhalothrin on aquatic invertebrates in laboratory and outdoor microcosm tests

The sensitivity of a range of freshwater lentic invertebrates to gamma-cyhalothrin (GCH), a single enantiomer of the synthetic pyrethroid lambda-cyhalothrin, was assessed in single species laboratory tests and an outdoor multi-species ecosystem test. The most sensitive species in the laboratory single species tests with GCH was Chaoborus obscuripes (96 h EC(50): 3.8 ng/l). The species sensitivity distribution curve, based on the laboratory 96 h EC(50) acute toxicity data for eight species, gave a median HC(5) value for GCH of 2.12 ng/l. The NOEC(community) derived from the multi-species ecosystem test was 5 ng/l, and the insects Chaoborus sp. and Caenis sp. were identified as the most sensitive species. The results indicate that the median HC(5), based on eight species selected to include those known to be sensitive to pyrethroids, provided a good estimation of the NOEC(community) for GCH. Furthermore, the results for GCH indicated that the endpoints typically used in higher-tier risk assessments for pesticides in Europe (HC(5) and NOEC(community)) were consistent with expectations when compared to the equivalent endpoints for the racemate LCH.

Effects of perfluorooctane sulfonate and perfluorooctanoic acid on the zooplanktonic community

This comparative survey summarizes six individual studies on the ecological effects of two common perfluorinated surfactants, PFOS and PFOA, on zooplankton. We compare the test designs and quantify the relative sensitivity and statistical power (1− β⩾0.8). The survey compares 30-L indoor microcosm to 12,000-L outdoor microcosm experiments, with 225-mL single species laboratory tests as reference. By this we elucidate the extrapolation of ecological effects in space and complexity. Generally, zooplankton had lower tolerance toward PFOS than toward PFOA. With increasing concentrations the zooplankton community became simplified toward more robust rotifer species, which, as an indirect effect, increased their abundance due to a shift in competition and predation. The statistical power of the designs exhibits inverse proportionality between complexity and realism, indoor microcosm>outdoor microcosm. Surprisingly, the 30-L study had a lower LOEC value for Daphnia magna than the laboratory chronic test, indicating that D. magna and D. pulicaria were not the most sensitive species and that laboratory tests are not always conservative relative to microcosm experiments. Food scarcity due to phytotoxicity was not the reason for the difference.

Organic contaminants in soil

For scientists, there is no shortcut to performing serious soil ecotoxicology. A fate description of the contaminants is included to explain effects; sometimes the effect assessment must be used to point to the presence of a chemical even though the fate is not clearly understood. Details of fate and effects are sometimes useless for risk assessment due to a lack of precision or a failure to link convincingly the fate to the effects or the biological parameters to the healthiness of the ecosystem. This is not a surprise in the single risk assessment case because limited resources preclude an iterative research strategy to reveal the mechanisms. Widespread risk assessment practices based on legislation, however, provide shortcuts by requiring an effect assessment derived from single-species laboratory tests. To derive predicted no-effect concentrations (PNECs) such tests have hitherto been used in connection with assessment factors and more recently with species sensitivity distribution models [1]. The latter assumes that the PNEC can be based on a complement of independent single-species toxicity data. During a conference held in Copenhagen Denmark that focussed on the fate and effects of organic contaminants in soil, it was shown that species interactions indeed are important for the toxicity levels of the individual species. Multispecies tests performed in a standardized manner are now slowly emerging, and they obviously include species interactions. Realism is gained at the cost of interpretability. This does not pose a problem, however, because interpretability is not paramount for risk assessment. There is still an important need to integrate the available information on the effects of all kinds of factors exerting environmental influences. Aging and bioavailability of the organic contaminants must be included and will greatly change the ecotoxicological setting of limit concentration in soil [2]. Subtle knowledge of chemical fate and effects is useless in the process of making sound environmental risk assessment if concomitant disturbances from other human or natural sources are not included. The risk assessment procedure would gain respect from all parties—academia, industry and environmental protection agencies—if it could be done in an integrated risk assessment process. Soil management in agriculture is a good example of an important factor whose impact can surmount effects of chemicals in many cases or change bioavailability in others. The reluctance to provide more environmental information is not rational; improved risk assessments will most probably ease the burden of administrating low-limit values and ensure that hazard is not overestimated. Variability in soil physicochemical properties represents a substantial scientific challenge to understanding the toxicology of soil contaminants. Bioavailability of organic soil contaminants became the driving theme of the Copenhagen meeting, and the role of this subject will have increased importance in the future. Scientists must focus on the general applicability of their results and consider the soil-aging factor as a major sink of persistent organic compounds. The political importance of this subject was clearly demonstrated: One leading Danish newspaper devoted its front page to quoting Martin Alexander on the role of aging in risk assessment of contaminated soils. In this issue of Environmental Toxicology and Chemistry you will find a selection of papers presenting current research on biodegradation and chemical availability.

Assessing the effects of environmental pollutants on soil organisms, communities, processes and ecosystems

A wide range of pollutants reach the soils of natural and managed ecosystems in concentrations that can affect their function. These chemicals, which include pesticides, heavy metals, acid deposition and a range of industrial chemicals, can reach soils in many different ways and by various routes. The ecological impacts of these chemicals on agricultural systems can involve effects at the: (i) organism population level, in terms of individual life histories (birth rate, numbers, growth, mortality); (ii) at the community level in terms of effects on plant/plant, plant/microbial, or plant/faunal interactions, species diversity and on soil food webs; (iii) at the ecosystem level, effects on primary and secondary productivity, organic matter breakdown and nutrient cycling; (iv) at the landscape level, changes in spatial heterogeneity of plants and soil organisms, material transfer of soil and nutrients, and hydrologic transfers of nutrients. Currently available methods of assessing the effect of pollutants include single species laboratory tests, a few multi-species assays, and integrated soil microcosms and terrestrial model ecosystems. The latter two methods produce data on the effects of pollutants on populations, communities and ecosystems as well as the fate of pollutants.

The Impact of a Copper Gradient on a Microarthropod Field Community

In order to evaluate the power of laboratory toxicity tests to predict the situation in the field, soil microarthropods were studied at the population and community levels in a copper contaminated field, and data were compared to the outcome of single species laboratory tests in the same soil type from a companion study. In addition to copper, the influence of several soil characteristics and plant cover data on soil microarthropod abundance and distribution was tested. Total microarthropod abundance was highest at intermediate copper concentrations, and linearly related to grass biomass. For single species populations no clear picture of abundance in relation to soil copper was seen. Two collembolan species, Folsomia quadrioculata and Folsomia fimetaria, were among the most sensitive. The Shannon-Wiener index of biodiversity decreased linearly with increasing soil copper concentrations. The application of multivariate statistics showed copper to be the parameter best fitting the distribution of microarthropod species in the field. A distinction could be made between sampling points with 50–199 mg Cu/kg soil and those with higher copper concentrations. The species composition of the microarthropod community was not correlated with plant species diversity or total grass biomass, but was affected by a nearby row of tall trees. Community analysis of field data proved as sensitive as laboratory testing, and seemed superior to single species field population data concerning detection of copper effects on soil microarthropods.

Inhibition of photosystem II (PSII) electron transport as a convenient endpoint to assess stress of the herbicide linuron on freshwater plants

Effects of the herbicide linuron on photosynthesis of the freshwater macrophytes Elodea nuttallii (Planchon) St. John, Myriophyllum spicatum L., Potamogeton crispus L., Ranunculus circinatus Sibth., Ceratophyllum demersum L. and Chara globularis (Thuill.), and of the alga Scenedesmus acutus Meyen, were assessed by measuring the efficiency of photosystem II electron flow using chlorophyll fluorescence. In a series of single-species laboratory tests several plant species were exposed to linuron at concentrations ranging from 0 to 1000 μg l−1. It was found that the primary effect of linuron, inhibition of photosystem II electron flow, occurred with a half-lifetime of about 0.1 to 1.9 h after addition of linuron to the growth medium. The direct effect of the herbicide on photosynthesis appeared to be reversible. Complete recovery from the inhibition occurred with a half-lifetime of 0.5 to 1.8 h after transfer of linuron treated plants to linuron free medium. The EC50,24h of the inhibition of photosystem II electron transport by linuron was about 9–13 μg l−1 for most of the macrophytes tested. For S. acutus the EC50,72h for inhibition of photosystem II electron flow was about 17 μg l−1 for the free suspension, and 22 μg l−1 for cells encapsulated in alginate beads. In a long-term indoor microcosm experiment, the photosystem II electron flow of the macrophytes E. nuttallii, C. demersum and the alga Spirogyra sp. was determined during 4 weeks of chronic exposure to linuron. The EC50,4weeks for the long-term exposure was 8.3, 8.7 and 25.1 μg l−1 for E. nuttallii, C. demersum and Spirogyra, respectively. These results are very similar to those calculated for the acute effects. The relative biomass increase of E. nuttallii in the microcosms was determined during 3 weeks of chronic exposure and was related to the efficiency of photosystem II electron transport as assessed in the different treatments. It is concluded that effects of the photosynthesis inhibiting herbicide on aquatic macrophytes, algae and algae encapsulated in alginate beads can be conveniently evaluated by measuring photosystem II electron transport by means of chlorophyll fluorescence. This method can be used as a rapid and non-destructive technique in aquatic ecological research.

The threshold problem in ecotoxicology

The most commonly used threshold in environmental toxicology is the LC50 (or modifications thereof) where 50% of the organisms die or are otherwise affected at a certain concentration of a chemical for a particular time of exposure under specified environmental conditions. Most commonly, this particular threshold is derived from single species laboratory tests low in environmental realism. If the field of ecotoxicology truly examines the effects of chemicals on ecosystems (i.e., complex multivariate systems), serious consideration must be given to thresholds other than those now commonly used in the field of environmental toxicology. Attributes at the community and ecosystem level of organization are not demonstrated at lower levels of biological organization, for example, energy flow and nutrient spiralling. Key issues are whether extrapolation is possible from one threshold to another within a level of biological organization and from one level of biological organization to another for thresholds that do not exist at many levels. Thresholds may be artefacts of testing procedures and may not exist in natural systems. Nevertheless, society must make management decisions about risk with available methods, including those designed to identify some point or threshold below which no deleterious effects are observed. However, these methods and their assumptions deserve more explicit and systematic examination than they have received thus far.

Incorporation of a subacute test with zebra fish into a hierarchical system for evaluating the effect of toxicants in the aquatic environment

Single-species laboratory tests were used to assess the acute toxicity of halogenated phenolic compounds. No single test system was most sensitive to all of the compounds examined, substantial variations in the sensitivity of the various organisms were noted, and there was no correlation between the toxicities assayed with different test systems. The zebra fish ( Brachydanio rerio) embryo/larvae system was used to examine subacute effects using two of the compounds, and a protocol was developed with 6 weeks preexposure to the toxicant. Preexposure decreased the lowest observable effect concentration by a factor of about 4, and the effect was completely reversible during a 6-week postexposure period in the absence of the toxicant. An enclosed system for carrying out the zebra fish embryo/larvae test was developed and evaluated with three neutral volatile compounds: the median survival time and the frequency of occurrence of deformation were examined as end points. The effect of pH on toxicity was evaluated in buffered media for four of the test systems: toxicity increased markedly at the lower pH values, and it could be shown that the ionized forms of the phenols were not the only contributors to toxicity. It is proposed that the zebra fish system incorporating preexposure could be incorporated into a hierarchical system using a range of organisms for assessing acute toxicity in single species under laboratory conditions and multicomponent systems simulating natural ecosystems.

Developing a statistical support system for environmental hazard evaluation

Estimating the hazard or risk to both human health and the environment has been based almost exclusively on single species toxicity tests low in environmental realism and without validation of their accuracy in more complex systems. While this may be quite appropriate for humans in a large variety of circumstances, there is no substantive body of direct experimental evidence indicating that precise predictions of harm from hazardous materials can be extrapolated from single species laboratory tests (or even multispecies laboratory tests) to the more complex highly variable natural systems. Now added to the hazardous chemical assessment problem is the accidental or deliberate release of genetically engineered microorganisms into the environment that have the additional capability of multiplying and expanding their numbers and also transferring genetic information to other organisms. This paper focuses entirely on hazard evaluation for organisms other than humans, namely predicting the potential risk or probability of harm to natural systems based on laboratory toxicity testing using single species. Not only will the basic risk assessment strategy itself be examined but also the question of determining the statistical reliability of various extrapolations from one level of biological organization to another.