Embryotoxicity of the alkylphenol degradation product 4-nonylphenol to the crustacean Daphnia magna

Laboratory studies have suggested that some alkylphenols and pesticides elicit developmental toxicity to crustaceans. The purpose of the present study was to evaluate the possibility that the alkylphenol degradation product 4-nonylphenol is embryotoxic to the crustacean Daphnia magna through its known ability to interfere with the metabolic elimination of testosterone. Direct exposure of maternal daphnids to testosterone caused developmental abnormalities in neonates that consisted of partial arrest of early embryonic development and abnormalities in shell spine and first antennae development. Exposure of maternal daphnids to concentrations of 4-nonylphenol also produced developmental abnormalities though the profile of abnormalities was distinct from that observed throughout the testosterone concentration-response curve. Thus, 4-nonylphenol is a developmental toxicant in daphnids, but its toxicity is not consistent with that elicited by elevated testosterone accumulation. Further experiments demonstrated that testosterone was directly toxic to developing embryos, and the maternal organism can serve as the vector for this toxicity. In contrast, neither direct embryo exposure nor early maternal exposure to 4-nonylphenol elicited embryotoxicity consistent with that observed during continuous maternal and gestational exposure. Thus, 4-nonylphenol is not directly embryotoxic at these exposure levels, but rather toxicity is mediated by maternal influences during gestation. The threshold concentration for the occurrence of developmental abnormalities ( approximately 44 microg/L) indicates that typical environmental concentrations of 4-nonylphenol pose no imminent hazard with respect to developmental toxicity. However, these effects do occur at sufficiently low levels to warrant evaluation of the relative susceptibility of other crustacean species to this previously uncharacterized mode of toxicity.     

Acute and chronic effects of testosterone and 4-hydroxyandrostenedione to the crustacean Daphnia magna

Steroid compounds have been globally detected in surface waters. The ecological impacts of these biologically active chemicals are largely unknown. Toxicity of testosterone and 4-hydroxyandrostenedione was assessed for the freshwater cladoceran Daphnia magna. Acute toxicity tests showed that 6.20 mg L⁻¹ of testosterone, the highest concentration tested, did not have effect on the daphnids, whereas 4-hydroxyandrostenedione had an EC₅₀ of 4.26 mg L⁻¹. Chronic toxicity tests were carried out using survival, body length, fecundity, and fertility as endpoints. Long-term testosterone exposure reduced D. magna fecundity and fertility at concentrations ranging from 0.31 to 2.48 mg L⁻¹. The significant decrease in fecundity was associated with an increase in aborted eggs. Long-term 4-hydroxyandrostenedione exposure at 0.84 mg L⁻¹ increased the mortality of the neonates. The chronic toxicity effects were observed at concentrations higher than the measured environmental concentrations of these compounds. Nevertheless, the reproductive impairment of the daphnids is likely to occur at environmental levels as an ultimate response to long-term exposure.     

The fungicide propiconazole interferes with embryonic development of the crustacean Daphnia magna

Propiconazole is a fungicide used in a variety of agricultural applications. Preliminary studies had suggested that embryos of the crustacean Daphnia magna are particularly susceptible to the toxicity of this chemical. The goals of the present study were to define endpoints of daphnid embryonic development that could be routinely used to assess the embryo toxicity of chemicals and to characterize definitively the embryo toxicity of propiconazole to daphnids. Daphnid embryonic development was characterized into six readily distinguishable stages based on the degree of tissue differentiation. Embryonic development could be monitored either in the brood chamber of the maternal organism or using embryos removed from the brood chamber and incubated ex vivo. Standard toxicity assessment revealed that propiconazole elicited no significant adverse effects on daphnid survival or fecundity during a 21-d exposure to concentrations as high as 0.25 mg/L. Exposure to 0.25 mg/L propiconazole, however, caused a significant incidence of developmental abnormalities and embryonic death. Abnormalities were consistent with developmental arrest at later stages of embryonic maturation. Propiconazole elicited a steep concentration-response curve with respect to embryo toxicity, with a 10% and a 90% incidence of embryo toxicity measured at 0.50 and 0.82 mg/L, respectively. Direct exposure of embryos to propiconazole resulted in toxicity, though the incidence and characteristics of developmental abnormalities were not consistent with that observed during chronic exposures. However, maternal exposure to propiconazole followed by transfer of early embryos to propiconazole-free media resulted in embryo toxicity consistent with that observed during chronic exposure. These results indicate that propiconazole interferes with the later stages of daphnid embryonic development, and that this toxicity is manifested largely via maternal exposure to the fungicide.     

Acute toxicity of the natural algicide, cyanobacterin, to Daphnia magna

Cyanobacterin is a potent, photosynthetic inhibitor produced as a secondary metabolite by the filamentous, freshwater cyanobacterium Scytonema hofmanni. With a spectrum of activity encompassing species of cyanobacteria and eucaryotic algae as well as higher plants, cyanobacterin might be utilized as a commercial algicide for algae waterbloom control. As a component of the preliminary environmental risk/hazard evaluation of this proposed application, the acute toxicity of cyanobacterin to the planktonic crustacean, Daphnia magna, was assessed. Based on five trials, the 48-hr LC50 for D. magna was determined to be approximately 1.37 micrograms/ml (range: 0.78-2.58 micrograms/ml). In addition, a significant percentage of the surviving daphnids were completely immobile at the end of the 48-hr exposure period. These results give strong indication that cyanobacterin and/or its residues exhibit some nontarget organism lethality at projected use-concentration level. Furthermore, estimates of partition coefficients and bioconcentration factor, generated through quantitative structure-activity relationships, suggest that sediment, suspended particulates, and biota may serve as major compartments of cyanobacterin partitioning in an aquatic ecosystem. Before cyanobacterin should be further considered for application as a commercial algicide, a rigorous evaluation of nontarget organism lethality and environmental fate is required.     

Effects of dehydroabietic acid and abietic acid on survival, reproduction, and growth of the crustacean Daphnia magna

Resin acids, a class of wood extractives, are potential toxic constituents in many pulp and paper mill effluents. In the present investigation, the effects of two predominant resin acids, dehydroabietic acid (DHA) and abietic acid (ABA), on survival, reproduction, and growth of the freshwater crustacean Daphnia magna were assessed over its life cycle. Based on the experimentally determined acute toxicity data (48-h EC(50)'s) for DHA (7.48 mg/L) and ABA (7.98 mg/L), D. magna was treated chronically with each resin acid at nominal concentrations of 0, 0.25, 0.5, 1.0, 2.0, 4.0, and 8.0mg/L for 21 days. Both DHA and ABA at concentrations as high as 4.0mg/L did not affect physiological and reproductive parameters such as time to maturation, number of molting, number of broods, and number of offspring produced from surviving daphnids, while significant mortality was observed only at 8.0mg/L in both cases. However, a small but statistically significant decrease in Daphnia growth (body length) at the end of exposure was detected at concentrations as low as 0.5mg/L for DHA and 1.0mg/L for ABA, respectively. These results indicated that both DHA and ABA had the potential to exhibit weak growth inhibition without apparent negative effects on reproduction to D. magna at nonlethal concentration levels. This slight effect is not expected to be ecologically significant because the concentrations of DHA and ABA in biologically treated pulp and paper mill effluents are well below the effective levels observed in the present study.     

Mixture toxicity of flubendazole and fenbendazole to Daphnia magna

Nowadays, residual amounts of many pharmaceuticals can be found in various environmental compartments including surface and ground waters, soils and sediments as well as biota. Even though they undergo degradability, their environmental discharge is relatively continuous, thus they may be regarded as quasi-persistent contaminants, and are also frequently regarded as emerging organic pollutants. Benzimidazoles, especially flubendazole (FLU) and fenbendazole (FEN), represent two anthelmintic drugs belonging to this group. Although their presence in environmental matrices has been reported, there is relatively little data concerning their (eco)toxicological impact. Furthermore, no data is available on their mixture toxicity. FLU and FEN have been found to have a strong impact on an environmentally important non-target organism – Daphnia magna. Moreover, these compounds are usually present in the environment as a part of pharmaceutical mixtures. Therefore, there is a need to evaluate their mixture toxicity, which was the main aim of this study. Single substance toxicity tests were carried out in parallel with mixture studies of FLU and FEN, with the application of two well established concepts of Concentration Addition (CA) and Independent Action (IA). As a result, both models (CA and IA) were found to underestimate the toxicity of mixtures, however CA yielded more accurate predictions.     

The effects of 4-nonylphenol and ethanol on acute toxicity,embryo development,and reproduction in Daphnia magna

The mean 48-h EC(50) (n=3) of 4-nonylphenol (NP) using ethanol as the carrier solvent was 155 microg/L, compared to a mean 48-h EC(50) (n=3) of 281 microg/L without ethanol. The 96-h EC(50)'s for embryo lethality (arrested egg development) and deformities (curved or unextended shell spines and undeveloped second antennae) were 738 and 263 microg/L, respectively. Reproduction studies were conducted using conditions that stimulate male production (i.e., reduced photoperiod and food levels). An increase in neonate deformities was observed at 50 microg/L (without ethanol), but no changes were observed in fecundity or sex ratios. A decrease in sex ratios was observed at 25 and 50 microg/L (with ethanol) compared to the ethanol control. However, an increase in sex ratios was observed in the ethanol control compared to media controls. The use of ethanol as a solvent carrier confounds the effects of 4-NP on acute toxicity and male production in daphnids.     

Influence of organism age on metal toxicity to Daphnia magna

Aquatic organisms living in surface water experience contaminant exposure at different life stages. While some investigators have examined the influence of organism age on the toxicity of pollutants, the general assumption in toxicology has been that young organisms were more sensitive than older organisms. In fact, some standardized toxicity tests call for the use of organisms less than 24 h old. This research characterized the age sensitivity of the water flea Daphnia magna to copper, zinc, selenium, and arsenic. During 21-d toxicity tests, organisms were exposed to a single 12-h pulse of either 70 microg/L Cu, 750 microg/L Zn, 1000 microg/L Se, or 5000 microg/L As at different ages ranging from 3 h to 10 d old. Mortality and reproduction were compiled over 21 d. During the juvenile stage, mortality increased and cumulative reproduction decreased with age, respectively. However, mortality decreased and cumulative reproduction increased with age when organisms became adult. Peak sensitivity occurred in 4-d-old organisms exposed to Cu and Zn, while 2- to 3-d-old organisms were most sensitive to As and Se. Growth of D. magna over 21 d was not affected by the 12-h pulse of Cu, Zn, Se, or As given at any organism age. This indicates the recovery of the organisms after exposure termination.     

Acute and chronic toxicity of imidazolium-based ionic liquids on Daphnia magna

Room-temperature ionic liquids (ILs) are considered to be green chemicals that may replace volatile organic solvents currently used by industry. However, IL effects on aquatic organisms and ecosystems are currently unknown. We studied the acute effects of imidazolium-based ILs on survival of the crustacean Daphnia magna and their chronic effects on number of first-brood neonates, total number of neonates, and average brood size. Lethal concentrations of imidazolium ILs with various anions (X-) ranged from a median lethal concentration (LC50) of 8.03 to 19.91 mg L(-1), whereas salts with a sodium cation (Na+ X-) were more than an order of magnitude higher (NaPF6 LC50, 9,344.81 mg L(-1); NaBF4 LC50, 4765.75 mg L(-1)). Thus, toxicity appeared to be related to the imidazolium cation and not to the various anions (e.g., Cl-, Br-, PF6-, and BF4-). The toxicity of imidazolium-based ILs is comparable to that of chemicals currently used in manufacturing and disinfection processes (e.g., ammonia and phenol), indicating that these green chemicals may be more harmful to aquatic organisms than current volatile organic solvents. We conducted 21-d chronic bioassays of individual D. magna exposed to nonlethal IL concentrations at constant food-resource levels. Daphnia magna produced significantly fewer total neonates, first-brood neonates, and average neonates when exposed to lower concentrations (0.3 mg L(-1)) of imidazolium-based ILs than in the presence of Na-based salts at higher concentrations (400 mg L(-1)). Such reductions in the reproductive output of Daphnia populations could cascade through natural freshwater ecosystems. The present study provides baseline information needed to assess the potential hazard that some ILs may pose should they be released into freshwater ecosystems.     

The uptake of cadmium from a dietary and soluble source by the crustacean Daphnia magna

Daphnia were exposed to radioactively labeled cadmium in solution and in the presence of Chlorella which had been preloaded with the metal to varying extents. Illuminated algal cells retained the cadmium and greatly reduced its availability to the daphnids. Autoradiographic evidence was obtained which implicated the exoskeleton as a major sink for the cadmium taken up from solution. Cadmium in solution at a concentration close to the 48 hr LC50 level did not affect respiration during the first 6 hr of exposure. Retention patterns were similar, regardless of the source of cadmium, but ecdysis resulted in a considerable loss of body burden provided that this had been acquired via a predominantly soluble route.     

Acute toxicity of furazolidone on Artemia salina, Daphnia magna, and Culex pipiens molestus larvae

As a result of evidence of the ecotoxicity of nitrofurans, the acute toxicity of furazolidone was tested in vivo on two aquatic organisms, Artemia salina and Daphnia magna, which are both crustaceans. Toxicity studies were also performed on larvae of Culex pipiens molestus. Results indicated a significant toxicity of the compound on Culex pipiens and Daphnia magna, while Artemia salina proved to be the least sensitive.     

Enantiospecific toxicity of the beta-blocker propranolol to Daphnia magna and Pimephales promelas

Propranolol is a widely prescribed, nonselective beta-adrenergic receptor-blocking agent. Propranolol has been detected in municipal effluents from the ng/L to the low-microg/L range. Like many therapeutics and other aquatic contaminants, propranolol is distributed as a racemic mixture ((R,S)-propranolol hydrochloride). Although the (S)-enantiomer is the most active form in mammals (up to 100-fold difference), no information is available regarding the enantiospecific toxicity of propranolol to aquatic organisms. Acute and chronic studies were conducted with Daphnia magna and Pimephales promelas to determine enantiospecific toxicity of propranolol to a model aquatic invertebrate and vertebrate, respectively. Also, enantiospecific effects of propranolol on D. magna heart rate were examined. Propranolol treatment levels were verified using high-performance liquid chromatography/mass spectrometry. Acute (48-h) responses of both organisms were similar for all enantiomer treatments. Chronic P. promelas responses to propranolol enantiomers followed the hypothesized relationship of (S)-propranolol being more toxic than (R)-propranolol, but chronic D. magna responses did not. This is potentially the result of a lack of beta-type receptors in cladocerans. No enantiospecific effects on daphnid heart rate were observed in acute exposures. Interestingly, some propranolol enantiomer treatments produced significant increases in reproduction before causing reproduction to decrease at higher treatment levels. To our knowledge, this research represents the first study of enantiospecific toxicity of chiral pharmaceutical pollutants.     

Insecticidal juvenile hormone analogs stimulate the production of male offspring in the crustacean Daphnia magna

Juvenile hormone analogs (JHAs) represent a class of insecticides that were designed specifically to disrupt endocrine-regulated processes relatively unique to insects. Recently we demonstrated that the crustacean juvenoid hormone methyl farnesoate programs oocytes of the crustacean Daphnia magna to develop into males. We hypothesized that insecticidal JHAs might mimic the action of methyl farnesoate, producing altered sex ratios of offspring. Daphnids were exposed chronically (3 weeks) to sublethal concentrations of methyl farnesoate, the JHA pyriproxyfen, and several nonjuvenoid chemicals to discern whether excess male offspring production is a generic response to stress or a specific response to juvenoid hormones. Only methyl farnesoate and pyriproxyfen increased the percentage of males produced by exposed maternal organisms. As previously reported with methyl farnesoate, acute exposure (24 hr) to either pyriproxyfen or the JHA methoprene caused oocytes maturing in the ovary to develop into males. We performed experiments to determine whether combined effects of a JHA and methyl farnesoate conformed better to a model of concentration addition (indicative of same mechanism of action) or independent joint action (indicative of different mechanisms of action). Combined effects conformed better to the concentration-addition model, although some synergy, of unknown etiology, was evident between the insecticides and the hormone. These experiments demonstrate that insecticidal JHAs mimic the action of the crustacean juvenoid hormone methyl farnesoate, resulting in the inappropriate production of male offspring. The occurrence of such an effect in the environment could have dire consequences on susceptible crustacean populations.     

A comparative study of rac- and S-metolachlor toxicity to Daphnia magna

Racemic product of metolachlor contains two R enantiomers and two S enantiomers. S-metolachlor is now widely used instead of rac-metolachlor because the former is more effective in herbicidal activity than the latter. The acute and chronic toxicities of rac- and S-metolachlor to Daphnia magna were determined and compared in this study. The acute 24-h LC50s for rac- and S-metolachlor to D. magna were 69.4 and 51.2 mgL(-1), respectively, indicating that the acute toxicity of S-metolachlor is slightly higher than that of rac-metolachlor. In chronic test, lowest-observed-effect concentration (LOEC), no-observed-effect concentration (NOEC), days to first brood, length, longevity, number of broods per female, number of young per female, and the intrinsic rate of natural increase (r) were determined. The results showed that the LOEC and NOEC of rac-metolachlor were 0.01 and 0.001 mgL(-1), while those of S-metolachlor were 0.5 and 0.1 mgL(-1), respectively. The first brood day of D. magna was not affected by rac- or S-metolachlor. Longevity and the number of broods per female were significantly (P<0.05) affected when rac-metolachlor concentration was higher than 1.0 mgL(-1) but were not significantly affected until S-metolachlor concentration was higher than 10 mgL(-1). The length was affected at the same concentration of rac- and S-metolachlor. Number of brood per female was significantly reduced when rac-metolachlor concentration was higher than 0.01 mgL(-1) but not significantly reduced until S-metolachlor concentration was higher than 0.5 mgL(-1). The r values were significantly reduced when the rac-metolachlor concentrations were above 0.01 mgL(-1) but not reduced only when S-metolachlor concentration was higher than 0.5 mgL(-1). The results of chronic toxicity indicated that rac-metolachlor was significantly (P<0.05) more toxic than S-metolachlor to D. magna.     

Maternal transfer efficiency and transgenerational toxicity of methylmercury in Daphnia magna

We examined maternal transfer efficiency, retention by subsequent generations, and transgenerational toxicity of methylmercury (CH3Hg or MeHg) in a population of freshwater zooplankton (Daphnia magna). The effect of dietary MeHg residence time in the daphnids on the efflux system also was quantified. After ingesting a relatively high dosage of MeHg, D. magna exhibited a reduction of live neonates and an increase of undeveloped eggs (or embryos), which reflected the sublethal toxicity of MeHg. The daily maternal transfer efficiency of MeHg to both reproductive outputs ranged from 0.42 to 4.9% over different ages of the parental daphnids, which was dependent on the daily reproductive output. During the lifetime of D. magna, reproduction contributed to 10.8% +/- 1.74% (n = 3) SD of total MeHg loss from the parental daphnids. The percentage of MeHg retention by the second generation (F1) of D. magna (40-60%) was generally higher than that by the parental generation (F0; approximately 25%) after 20 d of depuration. Methylmercury imposed sublethal toxicity to the F0 and F1 generations, but a smaller effect was observed on the F2 generation. Because of the very low MeHg body burden in the subsequent generations, we hypothesized that factors other than MeHg, such as nutritional deficiency in the offspring contributed to the transgenerational toxicity. Different MeHg residence times did not significantly affect the efflux rate of MeHg but did significantly affect the relative importance of reproduction as the elimination pathway for MeHg. Based on the MeHg body burden of neonates, we estimated that MeHg took 2.5 to 3.0 d to be optimally transferred from assimilation (e.g., gut) to the site of egg development (e.g., brood chamber) in D. magna. Our study demonstrated that maternal transfer of MeHg in freshwater zooplankton is an important predictor of MeHg concentration in their offspring and is a time-dependent and highly dynamic process.