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.

Bicarbonate alone does not totally explain the toxicity from major ions of coal bed derived waters to freshwater invertebrates.

Concentrations of major ions in coal mine discharge waters and unconventional hydrocarbon produced waters derived from coal bed methane (CBM) production, are potentially harmful to freshwater ecosystems. Bicarbonate is a major constituent of produced waters from CBM and coal mining. However, little is known about the relative toxicity of differing ionic proportions, especially bicarbonate, found in these CBM waters. As all freshwater invertebrates tested are more acutely sensitive to sodium bicarbonate (NaHCO3) than sodium chloride (NaCl) or synthetic sea water, we tested the hypotheses that toxicity of CBM waters are driven by bicarbonate concentration, and waters containing a higher proportion of bicarbonate are more toxic to freshwater invertebrates than those with less bicarbonate. We compared the acute (96 h) lethal toxicity to six freshwater invertebrate species of NaHCO3 and two synthetic CBM waters, with ionic proportions representative of water from CBM wells across New South Wales (NSW) and Queensland (Qld), in Australia. The ranking of LC50 values expressed as total salinity was consistent with the hypotheses. However, when toxicity was expressed as bicarbonate concentration, the hypothesis that the toxicity of coal bed waters would be explained by bicarbonate concentration was not well supported, and other ionic components were either ameliorating or exacerbating the NaHCO3 toxicity. Our findings showed NaHCO3 was more toxic than NaCl and that the NaHCO3 proportion of synthetic CBM waters drives toxicity, however other ions are altering the toxicity of bicarbonate.

Species of freshwater invertebrates that are sensitive to one saline water are mostly sensitive to another saline water but an exception exists.

Coal mining and extraction of methane from coal beds generate effluent with elevated salinity or major ion concentrations. If discharged to freshwater systems, these effluents may have adverse environmental effects. There is a growing body of work on freshwater invertebrates that indicates variation in the proportion of major ions can be more important than salinity when determining toxicity. However, it is not known if saline toxicity in a subset of species is representative of toxicity across all freshwater invertebrates. If patterns derived from a subset of species are representative of all freshwater invertebrates, then we would expect a correlation in the relative sensitivity of these species to multiple saline waters. Here, we determine if there is a correlation between the acute (96 h) lethal toxicity in freshwater invertebrates to synthetic marine salts (SMS) and sodium bicarbonate (NaHCO3) added to dechlorinated Sydney tap water. NaHCO3 is a major component of many coal bed effluents. However, most salinization in Australia exhibits ionic composition similar to seawater, which has very little HCO3- Across all eight species tested, NaHCO3 was 2-50 times more toxic than SMS. We also observed strong correlations in the acute toxicity of seven of the tested species to SMS and NaHCO3 The strongest relationship (LC50 r2 = 0.906) was dependent on the exclusion of one species, Paratya australiensis (Decopoda: Atyidae), which was the most sensitive species tested to NaHCO3, but the second-most tolerant of SMS. We conclude that differences in the toxicity of different proportions of major ions can be similar across a wide range of species. Therefore, a small subset of the invertebrate community can be representative of the whole. However, there are some species, which based on the species tested in the current study appear to be a minority, that respond differently to saline effluent and need to be considered separately. We discuss the implications of this study for the management of saline coal bed waters.This article is part of the theme issue 'Salt in freshwaters: causes, ecological consequences and future prospects'.

Bicarbonate toxicity to Ceriodaphnia dubia and the freshwater shrimp Paratya australiensis and its influence on zinc toxicity.

Bicarbonate is often a major ionic constituent associated with produced waters from methane gas extraction and coal mining, yet few studies have determined its specific toxicity. Currently, the environmental risk of bicarbonate anion in water discharges is assessed based on the toxicity of sodium chloride or artificial seawater and is regulated via electrical conductivity. Increased NaHCO(3) added to Ceriodaphnia dubia in synthetic or natural water gave similar 48-h 10% effective concentration (EC10) values of 1750 ± 125 mg NaHCO(3)/L (mean ± standard error) and 1670 ± 180 mg NaHCO(3)/L, respectively. Bicarbonate was toxic to C. dubia in both waters with conductivities above 1900 µS/cm. In contrast, when conductivity was elevated with NaCl, toxicity to C. dubia was observed only above 2800 µS/cm. Bicarbonate also impaired C. dubia reproduction with an EC10 of 340 mg NaHCO(3)/L. Major ion composition also influenced Zn bioavailability, a common co-occurring metal contaminant in coal mine waters, with sublethal concentrations of NaHCO(3) and elevated pH increasing Zn toxicity. Higher pH was the dominant parameter determining a 10-fold increase in the 48-h 50% effective concentration (EC50) for Zn toxicity to C. dubia at pH 8.6 of 34 µg Zn/L (95% confidence limit = 32-37 µg Zn/L) compared with the Zn toxicity at approximately circumneutral pH. Exposure of the freshwater shrimp Paratya australiensis (Atyidae) in natural water to increasing bicarbonate gave a mean 10-d 10% lethal concentration (LC10) of 850 ± 115 mg NaHCO(3)/L, associated with a mean conductivity EC10 of 1145 µS/cm, which is considerably lower than toxicity of NaCl and artificial seawater to this species reported elsewhere. Because toxicity was influenced by salt composition, specific ions should be regulated rather than conductivity alone in mine wastewater discharges.

Detection and analysis of neonicotinoids in river waters--development of a passive sampler for three commonly used insecticides.

Increasing and widespread use of neonicotinoid insecticides all over the world, together with their environmental persistence mean that surface and ground waters need to be monitored regularly for their residues. However, current multi-residue analytical methods for waters are inadequate for trace residue analysis of these compounds, while passive sampling devices are unavailable. A new method using UltraPerformance Liquid Chromatography provided good separation of the five most common neonicotinoid compounds, with limits of quantitation in the range 0.6-1.0ng. The method was tested in a survey of rivers around Sydney (Australia), where 93% of samples contained two or more neonicotinoids in the range 0.06-4.5µgL(-1). Styrenedivinylbenzene-reverse phase sulfonated Empore™ disks were selected as the best matrix for use in passive samplers. Uptake of clothianidin, imidacloprid and thiacloprid in a flow-through laboratory system for 3weeks was linear and proportional to their water concentrations over the range 1-10µgL(-1). Sampling rates of 8-15mLd(-1) were correlated to the hydrophobicity of the individual compounds. The passive samplers and analytical methods presented here can detect trace concentrations of neonicotinoids in water.

Differential survival and reproductive performance across three mitochondrial lineages in Melita plumulosa following naphthalene exposure.

Populations subject to anthropogenic contaminants often display altered patterns of genetic variation, including decreased genetic variability. Selective pressures of contaminant exposure are also reflected in differential tolerance between genotypes. An industrial chemical spill in a major eastern Australian waterway in July 2006 resulted in altered patterns of genetic variability in a nearby population of the amphipod, Melita plumulosa for up to one year post-spill, despite the site being declared clean after 48 h. Here, we investigate the toxicant response of three mitochondrial lines naturally occurring at the impacted site by comparing survivorship and life-history trait variables following naphthalene exposure. Overall, M. plumulosa demonstrated differential survivorship between mitochondrial lines under exposure to high concentrations of naphthalene. In addition, we identified differential fecundity and frequencies of gravidity in female amphipods between the mitochondrial haplotypes examined. These findings suggest that the patterns of genetic variability previously identified may be linked with differential tolerance and/or reproductive performance between mitochondrial lineages.

Secondary vitellogenesis persists despite disrupted fecundity in amphipods maintained on metal-contaminated sediment: X-ray fluorescence assessment of oocyte metal content.

Melita plumulosa is an epibenthic, detritivorous amphipod found in estuaries along the eastern coast of Australia. It has been utilized as a test organism in rapid ten to thirteen days reproduction toxicity tests for sediment quality assessment. The fecundity of females in the toxicity test has been found to be inhibited by exposure of the amphipods to contaminated sediments enriched with zinc and other metals. This study investigated the proposal that interference in vitellogenesis is the cause of reproductive toxicity of metals in crustaceans. Inspection of the ovaries from amphipods on day 6 of the test either from control or Zn/Pb/Cd/Cu-spiked sediment, that were nearing completion of vitellogenesis, showed that the females in all treatments were producing similar numbers of oocytes undergoing secondary vitellogenesis. The distribution of the Zn, Cu and Pb in the oocytes and ventral caeca of females was examined by X-ray fluorescence microscopy. Elemental mapping revealed a dense accumulation of Zn in primary oocytes and a uniform distribution of Zn and Cu in the secondary oocytes in all treatments. Zn and Cu were also observed to be uniformly distributed in the ventral caeca. Pb was not detected in either of these tissues. The apparent normal morphology and the typical number of oocytes undergoing secondary vitellogenesis suggest that vitellogenesis was not being disrupted by Pb displacing Zn in the metal-binding domain of vitellogenin in amphipods exposed to the contaminated sediment during the test. Alternative mechanisms for the reproductive toxicity of amphipods exposed for six days to metal-contaminated sediment are discussed.

Calibration and field application of Chemcatcher® passive samplers for detecting amitrole residues in agricultural drain waters.

A passive sampler device suitable for monitoring of residues of the hydrophilic ionic herbicide amitrole in irrigation waterways was developed. Uptake of amitrole on styrenedivinylbenzene-reverse phase sulfonated Empore™ disks was linear and proportional to its water concentration over the range of 1-10 µg/L with a sampling rate of 23.1 mL/day under laboratory flow-through conditions. Performance of the sampler was evaluated by deployment in an agricultural irrigation drain for 10 days. The amount of amitrole adsorbed by the passive samplers compared well with the cumulative mean water concentrations calculated from daily spot samplings of the drain water.

The ecological effects of a herbicide-insecticide mixture on an experimental freshwater ecosystem.

The effects of a co-occurring insecticide-herbicide mixture were evaluated using model ecosystems (microcosms) in the laboratory. Microcosms dosed with a high concentration (10 µg/L) of the insecticide terbufos, alone and as a mixture with the herbicide atrazine (25 µg/L), exhibited community level effects attributed to the elimination and decline of invertebrate populations, and also indirect effects. There were no community level effects at a lower concentration of terbufos (0.1 µg/L) alone or in a mixture with atrazine, although delayed population effects were observed. Female chironomids also emerged later and those from terbufos-only microcosms were smaller. Exposure to atrazine alone was also associated with lower abundances of cladocerans and reduced emergence of chironomids. The risk posed by atrazine is low and is unlikely to exacerbate the effects of terbufos. Nevertheless, the population-level effects highlight that terbufos poses a potential risk to aquatic ecosystems, regardless of whether atrazine is also present.

Review of the reproductive biology of amphipods and their endocrine regulation: identification of mechanistic pathways for reproductive toxicants.

The reproductive biology of amphipods is reviewed to update the knowledge of the male and female reproductive processes of oogenesis and spermatogenesis as well as the endocrine systems of amphipods with the aim of advancing studies of reproductive toxicology. The ovarian and reproduction cycles of female gammaridean amphipods are closely correlated with the molt cycle, which is under direct control by the steroid hormone 20-hydroxyecdysone. The ability of males to copulate and subsequently for females to ovulate is restricted to the early postmolt period of the females. New developments in our understanding of the molt cycle and the endocrine regulatory pathways for reproduction using genomics techniques on other crustacean species are also discussed. The arthropod sterol ponasterone A or xenobiotics such as the fungicide fenarimol have been shown to elicit endocrine disruption in some crustaceans by acting as an agonist for 20-hydroxyecdysone at the ecdysone receptor or by inhibiting the synthesis of 20-hydroxyecdysone, respectively, resulting in disruption of molting and reproduction. Recent studies suggest that cadmium can inhibit secondary vitellogenesis in amphipods. Experimental approaches for examining the metabolic pathways associated with ecdysteroid hormonal signaling or metabolism, exoskeleton maintenance and molting, and the regulation of vitellogenin in amphipods are discussed. This information should aid in the identification of useful biomarkers for reproductive toxicity.

Developmental toxicity of two common corn pesticides to the endangered southern bell frog (Litoria raniformis).

To examine the link between corn agriculture and the observed decline of the endangered southern bell frog (SBF), the effects of two corn crop pesticides on larval growth and development were investigated. Tadpoles were exposed to terbufos sulfone (10 µg/L), a major breakdown product of the insecticide terbufos, and the herbicide atrazine (25 µg/L) individually and as a mixture until the completion of metamorphosis. Atrazine did not interact synergistically with terbufos sulfone or result in significant effects on growth and development alone, although there was some indication of accelerated metamorphosis in the pilot study. Terbufos sulfone alone and as a mixture (terbufos/atrazine) significantly slowed larval development and ultimately delayed metamorphosis. The observed developmental effects from an environmentally relevant concentration of terbufos sulfone indicates a risk posed by this persistent degradation product to the endangered SBF, which breeds and develops in the rice bays adjacent to corn fields treated with pesticides.

Foraging, feeding, and reproduction on silica substrate: Increased waterborne zinc toxicity to the estuarine epibenthic amphipod Melita plumulosa.

Artificial substrates consisting of fine milled silica with or without α-cellulose were evaluated for their capacity to support survival, growth, and fecundity in the amphipod Melita plumulosa. There were no significant differences in the survival and fecundity of adult amphipods maintained for up to 13 d on natural sediment, silica-only, or silica/α-cellulose substrate when fed two algal foods, Sera micron and Rotiselco-ALG. However, growth among juveniles maintained on the silica/α-cellulose mixture was significantly inhibited over 14 d compared with natural sediment. Addition of a microencapsulated shrimp feed, Frippak, to the algal foods improved juvenile growth over 28 d but still did not match the nutritive value of natural sediment. Fine silica without cellulose was subsequently used in acute and reproductive toxicity tests with waterborne zinc. With food, a 10-d median lethal concentration (LC50) of 140 µg Zn/L and a 10-d no-effect concentration (NEC) of 80 µg Zn/L were obtained for juvenile survival on silica. In contrast, a 10-d LC50 of 200 µg Zn/L and a 10-d NEC of 180 µg Zn/L were obtained for juveniles in water-only exposures. Similarly, exposure of adult females to Zn without food on silica compared with water-only exposures gave 10-d LC50s of 380 and 490 µg Zn/L and 10-d NECs of 130 and 370 µg Zn/L, respectively. The reproduction toxicity test indicated significant adult mortality at 92 µg Zn/L and significantly reduced fecundity at 22 µg Zn/L. We surmised that the toxicity of waterborne zinc to M. plumulosa increased when maintained on nutrient-depleted silica compared with water-only exposure because of increased energy expended through foraging, in concert with the likely increased exposure to Zn via the digestive tract and the gills.

Comparison of environmental risks of pesticides between tropical and nontropical regions.

A comparison of environmental risks of pesticides between tropical and nontropical regions has been performed, using data from the literature and modeling outputs based on the physicochemical properties of the compounds. With a few exceptions, the level of risk of exposure for most pesticides in tropical agriculture is similar to that in other climatic regions of the world. Generally, dissipation of pesticides increases under the warm and wet conditions of the tropics, with most of the dissipation occurring through hydrolysis in water and biological degradation in water and soil. High temperatures in the tropics also foster volatilization rates, whereas high precipitation and poor soils tend to increase losses into runoff and, for certain chemicals, affects their leaching behavior. The environmental risk is determined by a balance of soil types, soil organic carbon, pH, and the rates of degradation in the various environmental compartments.

Temporal and geographical genetic variation in the amphipod Melita plumulosa (Crustacea: Melitidae): Link of a localized change in haplotype frequencies to a chemical spill.

Anthropogenic effects such as contamination affect the genetic structure of populations. This study examined the temporal and geographical patterns of genetic diversity among populations of the benthic crustacean amphipod Melita plumulosa in the Parramatta River (Sydney, Australia), following an industrial chemical spill. The spill of an acrylate/methacrylate co-polymer in naphtha solvent occurred in July 2006. M. plumulosa were sampled temporally between December 2006 and November 2009 and spatially in November 2009. Genetic variation was examined at the mitochondrial cytochrome c oxidase subunit I locus. Notably, nucleotide diversity was low and Tajima's D was significantly negative amongst amphipods collected immediately downstream from the spill for 10 months. We hypothesize that the spill had a significant localized effect on the genetic diversity of M. plumulosa. Alternate explanations include an alternate and unknown toxicant or a localized sampling bias. Future proposed studies will dissect these alternatives.

Toxicity of the insecticide terbufos, its oxidation metabolites, and the herbicide atrazine in binary mixtures to Ceriodaphnia cf dubia.

The acute toxicity of terbufos and its major metabolites, tested alone, in binary mixtures or in combination with atrazine were evaluated using neonates of the cladoceran Ceriodaphnia cf dubia. Terbufos, terbufos sulfoxide, and terbufos sulfone tested individually were highly toxic to C. cf dubia, with mean 96-h EC(50) values of 0.08, 0.36, and 0.19 µg/l, respectively. The addition of atrazine (10 µg/l) significantly increased the toxicity of terbufos. The toxicity of terbufos sulfone was unaffected by atrazine, whereas the results for terbufos sulfoxide were equivocal. Equitoxic mixtures of the metabolites showed additive toxicity to C. cf dubia. The high toxicities of terbufos and its environmentally persistent oxidative metabolites suggest that contamination of aquatic systems with this insecticide mixture and the coapplied herbicide atrazine might pose a greater hazard to some biota than their individual toxicities.

Morphological abnormalities in frogs from a rice-growing region in NSW, Australia, with investigations into pesticide exposure.

Three frog species (Limnodynastes tasmaniensis, Limnodynastes fletcheri and Litoria raniformis) were surveyed in rice bays of the Coleambally Irrigation Area (CIA), NSW, Australia, during the rice-growing seasons of 2005/2006 and 2006/2007. A total external morphological abnormality index of 7.0% was observed in frogs of the CIA (n=1,209). The types and frequencies of abnormalities were typical of reports from agricultural areas with ectrodactyly being the most common aberration. A relatively low abnormality index of 1.2% was observed in L. raniformis (n=87) compared to indices of 7.1% and 8.2% observed in L. fletcheri (n=694) and L. tasmaniensis (n=428), respectively. No conclusive evidence was found of unnaturally high rates of intersex, gonadal maldevelopment or unbalanced sex ratios in any species. Rice bay surface waters differed significantly in mean pesticide concentrations of atrazine and metolachlor on farms growing rice and corn compared to farms with rice as the sole crop. However, the similar abnormality indices observed in recent metamorphs emerging from these two farm types provided no evidence to suggest a link between larval exposure to the measured pesticides and developmental malformations.

A rapid amphipod reproduction test for sediment quality assessment: In situ bioassays do not replicate laboratory bioassays.

An underlying assumption of laboratory-based toxicity tests is that the sensitivity and exposure of organisms in the laboratory is comparable to that in the field. We sought to make a comparison between field-based and laboratory-based sediment toxicity tests using a recently developed rapid amphipod reproduction test that encompasses gametogenesis, fertilization, and embryo development before hatching. The test species, Melita plumulosa, is an epibenthic, detritivorous amphipod native to Eastern Australia. Test sediments were sourced from Lake Macquarie, a large saltwater lagoon located 100 km north of Sydney (New South Wales, Australia) that has received heavy-metal pollution over many decades, primarily from a Pb/Zn smelter but also from collieries, coal-fired power stations, and urban areas. This has led to a north-south trace-metal concentration gradient, including Pb, Zn, Cd and Cu, in the sediments of Lake Macquarie. Sediments from these northern bays were demonstrated to reduce amphipod fecundity in laboratory-based tests. For the current study, the amphipod reproduction test has been modified for use in situ. In situ test chambers were deployed at the mouth of Cockle Creek, Lake Macquarie. Sediments that were demonstrated to reduce fecundity of M. plumulosa in the laboratory reproduction test were not similarly toxic when amphipods were exposed to the same sediments in situ. Factors related to the regular tidal renewal of overlying water likely altered exposure profiles in situ, including the provision of additional or alternative nutrition that obviated the need for amphipods to interact with the contaminated sediments, and a washout effect that prevented the accretion of dissolved zinc in the overlying water.

Effects of dietary cadmium exposure on reproduction of saltwater cladoceran Moina monogolica Daday: Implications in water quality criteria.

The chronic toxicity of dietary cadmium to the saltwater cladoceran Moina monogolica Daday and its relative toxicity compared with aquatic exposure were investigated in the present study. The microalgae Chlorella pyrenoidosa, exposed to cadmium in growth inhibition tests, had a 96-h median effective concentration (EC50) of 81.2 microg Cd/L (95% confidence intervals [CIs] = 71.9-95.1). C. pyrenoidosa exposed for 96 h to sublethal dissolved cadmium concentrations in the range 4.10 +/- 0.30 to 70.29 +/- 0.31 microg/L resulted in algal cadmium burdens up to 73.86 x 10(-16) g Cd/cell. Cellular cadmium burdens accumulated in a dose-dependent manner, whereas cell densities inversely declined from 670 x 10(4) to 38 x 10(4) cells/ml with exposure to the increasing aqueous cadmium concentrations. C. pyrenoidosa preexposed to cadmium and used as food in a chronic 21-d toxicity test with the cladoceran M. monogolica, containing no added dissolved cadmium, inhibited reproduction. Significant reductions of the net reproduction rate (R(0)) per brood were observed in all broods, and the decline in the number of neonates produced increased with each subsequent brood. The cadmium concentration (4.10 +/- 0.30 microg/L) in the algal culture water that produced the lowest algal cadmium burden (2.85 +/- 0.76 x 10(-16) g Cd/cell) was shown to inhibit M. monogolica reproduction and was compared with the water quality criteria (WQC) of China. This comparison indicated that dietary exposure to cadmium may cause sublethal responses at concentrations below the current cadmium WQC of China for aquaculture.

An amperometric method for the detection of amitrole, glyphosate and its aminomethyl-phosphonic acid metabolite in environmental waters using passive samplers.

The herbicides amitrole and glyphosate, and its metabolite aminomethyl-phosphonic acid (AMPA), in water samples have been directly analysed by high-performance liquid chromatography using an electrochemical (EC) detector. Limits of detection of 0.3 microg mL(-1) for glyphosate, 0.05 microg mL(-1) for AMPA and 0.03 microg mL(-1) for amitrole were comparable to those obtained by other authors using EC and also by liquid chromatography coupled to mass spectrometry, but the latter method requires derivatisation and pre-concentration of the sample whereas EC methods show similar sensitivity without the need of any derivatisation. The method was specifically designed to analyse extracts from passive samplers used for monitoring of polar herbicide residues in waters. To this purpose, three types of Empore disks were tested for their ability to adsorb and desorb these ionic, polar analytes. A procedure for their extraction from the membranes and reducing the interferences from other substances present in natural waters (i.e. humic acids) is described. The method is simple, does not require sophisticated equipment and is valid for the analysis and monitoring of herbicides residues using passive samplers.

A low concentration of atrazine does not influence the acute toxicity of the insecticide terbufos or its breakdown products to Chironomus tepperi.

The acute toxicities of the insecticide terbufos and its major breakdown products individually, as binary mixtures, and in combination with the co-applied herbicide atrazine were evaluated using final instar larvae of the midge Chironomus tepperi. Terbufos, terbufos sulfoxide and terbufos sulfone were highly toxic to C. tepperi with mean 96-h EC50 values of 2.13, 3.64 and 2.59 µg/l, respectively. No interaction was observed between atrazine (25 µg/l) and terbufos or its breakdown products while the binary mixture of terbufos sulfoxide and terbufos sulfone exhibited additive toxicity. The high toxicities of terbufos and its environmentally persistent oxidation products suggest that contamination of aquatic systems with this insecticide pose a threat to aquatic organisms whether or not atrazine is also present.