Chronic toxicity of parabens and their chlorinated by‐products in Ceriodaphnia dubia

The chronic toxicity of 12 compounds of parabens and their chlorinated by‐products was investigated using 7‐day Ceriodaphnia dubia test under static renewal condition in order to generate information on how to disinfect by‐products of preservatives that are discharged in aquatic systems. The mortality and inhibition of reproduction tended to increase with increasing hydrophobicity and decreased with the degree of chlorination of parabens. The EC₅₀ values for mortality, offspring number, and first brood production ranged between 0.30–3.1, 0.047–12, and 1.3–6.3 mg L^?1, respectively. For the number of neonates, the most sensitive endpoint, the no‐observed‐effect concentration (NOEC) and lowest‐observed‐effect concentration (LOEC) values ranged from 0.63 to 10 mg L^?1 and from 1.2 to 19 mg L^?1, respectively. Methylparaben (MP), benzylparaben (BnP), and dichlorinated BnP (Cl₂BnP) elicited a significant decrease in offspring numbers even at their lowest concentration tested; the NOEC for these compounds was determined to be less than the lowest test concentration (1.3, 0.04, and 0.63 mg L^?1 for MP, BnP, and Cl₂BnP, respectively). Propylparaben (PP), chlorinated PP, isopropylparaben (iPP), and chlorinated iPP exhibited nonmonotonic concentration‐dependent response; their NOEC and LOEC values could not be determined. The multivariate approach involving principal component analysis and hierarchical cluster analysis revealed four groups that corresponded to the toxicological profiles of parabens. Our results suggested that disinfection of parabens by chlorination could reduce aquatic toxicity of original compounds. The findings obtained in our study together with the data available on paraben concentrations in aquatic systems can be used to perform preliminary risk assessment by comparing the predicted environmental concentration (PEC) with the predicted no‐effect concentration (PNEC) for the marine aquatic environment. The calculated PEC/PNEC ratios ranged from 0.0012 to 0.2, with the highest value observed in MP. This suggested that there are negligible environmental risks for aquatic organisms at current use levels. © 2013 Wiley Periodicals, Inc. Environ Toxicol 30: 664–673, 2015.     

Toxicological effects of military fog oil obscurant on Daphnia magna and Ceriodaphnia dubia in field and laboratory exposures

Our purpose was to determine if the acute and sub-lethal effects of fog oil, an obscurant used for military training, could be observed in realistic field exposures. To this end, we exposed Daphnia magna to oil fogs under actual release conditions at a U.S. Army training site. Guided by field investigations, acute toxicity experiments were conducted in the laboratory with the more sensitive species Ceriodaphnia dubia to test the hypothesis that dissolution of fog oil constituents into water is minimal and actual contact by organisms with the water surface is required to cause toxicity. We conducted further experiments to test the hypothesis that vaporization of fog oil alters its chemical composition and toxicity to freshwater invertebrates. In the field, daphnid mortality was minimal more than 5 m from the point of fog generation, but sub-lethal effects were more extensive. Both field and laboratory experiments suggested that physical contact with oils on the water surface was the most important factor driving toxicity. To our knowledge, this is the first attempt to evaluate toxicological endpoints with freshwater invertebrates in field exposures with fog oil.     

Chronic toxicity of silver nitrate to Ceriodaphnia dubia and Daphnia magna, and potential mitigating factors

We investigated the chronic toxicity of Ag, as silver nitrate, using two freshwater aquatic cladoceran species, Ceriodaphnia dubia and Daphnia magna, to generate data for the development of a chronic ambient water quality criterion for Ag. Preliminary studies with C. dubia showed variable results which were related to the equilibration time between food and silver. Follow-up testing was conducted using a 3h equilibration time, which stabilized dissolved Ag concentrations and the toxicity of Ag(+). Results with C. dubia conducted individually (1 per cup, n=10) and in mass (30 per chamber, n=2) gave similar results once similar standardized equilibration times were used. The maximum acceptable toxicant concentration (MATC) of Ag to C. dubia and D. magna was 9.61 and 3.00microg dissolved Ag/L, respectively. The chronic toxicity of Ag(+) to C. dubia was also evaluated in the presence of: (1) dissolved organic carbon (DOC) and (2) sulfide. The addition of DOC (0.4mg/L) resulted in a approximately 50% decrease in toxicity while the addition of sulfide (75.4nM) deceased toxicity by 42%. Whole-body Ag concentration in D. magna was positively correlated with increased levels of Ag exposure, however; we observed a non-statistical decrease in whole-body Na levels, an estimator of sodium homeostasis.     

Acute toxicity of 50 metals to Daphnia magna

Metals are essential for human life and physiological functions but may sometimes cause disorders. Therefore, we conducted acute toxicity testing of 50 metals in Daphnia magna: EC50s of seven elements (Be, Cu, Ag, Cd, Os, Au and Hg) were < 100 µg l^?1; EC50s of 13 elements (Al, Sc, Cr, Co, Ni, Zn, Se, Rb, Y, Rh, Pt, Tl and Pb) were between 100 and 1000 µg l^?1; EC50s of 14 elements (Li, V, Mn, Fe, Ge, As, In, Sn, Sb, Te, Cs, Ba, W and Ir) were between 1,001 and 100,000 µg l^?1; EC50s of six elements (Na, Mg, K, Ca, Sr and Mo) were > 100,000 µg l^?1; and. 7 elements (Ti, Zr, Bi, Nb, Hf, Re and Ta) did not show EC50 at the upper limit of respective aqueous solubility, and EC50s were not obtained. Ga, Ru and Pd adhered to the body of D. magna and physically retarded the movement of D. magna. These metals formed hydroxides after adjusting the pH. Therefore, here, we distinguished this physical effect from the physiological toxic effect. The acute toxicity results of 40 elements obtained in this study were not correlated with electronegativity. Similarly, the acute toxicity results of metals including the rare metals were also not correlated with first ionization energy, atomic weight, atomic number, covalent radius, atomic radius or ionic radius. Copyright © 2014 John Wiley & Sons, Ltd.     

Comparison of acute to chronic ratios between silver and gold nanoparticles,using Ceriodaphnia dubia

As integration of nanoparticles (NPs) into products becomes more common, the need to address the paucity of chronic hazard information for aquatic environments required to determine risk potential increases. This study generated acute and chronic toxicity reference values for Ceriodaphnia dubia exposed to 20 and 100?nm silver nanoparticles (AgNPs) and gold nanoparticles (AuNPs) to generate and evaluate potential differences in acute-to-chronic ratios (ACR) using two different feeding methods. A modified feeding procedure was employed alongside the standard procedures to investigate the influence of food on organism exposure. An 8-h period before food was added allowed direct organism exposure to NP dispersions (and associated ions) without food-to-NP interactions. The AgNPs [chronic lethal median concentrations (LC50) between 18.7 and 31.9?µg/L] were substantially more toxic than AuNPs (LC50?=?21 507 to >26 384?µg/L). The modified chronic testing method resulted in greater sensitivity in AgNPs exposures. However, the modified feeding ration had less of an effect in exposures to the larger (100?nm) AgNPs compared to smaller particles (20?nm). The ACRs for AgNPs using the standard feeding ration were 1.6 and 3.5 for 20?nm and 100?nm, respectively. The ACRs for AgNPs using the modified feeding ration were 3.4 and 7.6 for 20?nm and 100?nm NPs, respectively. This supports that the addition of the standard feeding ration decreases C. dubia chronic sensitivity to AgNPs, although it must also be recognized organisms may be sensitized due to less access to food. The ACRs for 20?nm and 100?nm AuNPs (standard ration only) were 4.0 and 3.0, respectively. It is important to also consider that dissolved Ag⁺ ions are more toxic than AgNPs, based on both acute toxicity values in the cited literature and chronic toxicity thresholds generated in this study that support existing thresholds that Ag⁺ are likely protective of AgNPs effects.     

Multi-level toxicity assessment of engineered cellulose nanofibrils in Daphnia magna

Cellulose nanofibril (CNF)-based materials are increasingly used in industrial and commercial applications. However, the impacts of CNF on aquatic life are poorly understood, and there are concerns regarding their potential toxicity. Using a combination of standard ecotoxicological tests and feeding experiments, we assessed the effects of CNF exposure (0.206–20.6?mg/L) on the feeding (food uptake and gut residence time) and life-history traits (growth and reproduction) in the cladoceran Daphnia magna. No mortality was observed in a 48?h acute exposure at 2060?mg/L. Moreover, a 21-day exposure at low food and moderate CNF levels induced a stimulatory effect on growth, likely driven by increased filtration efficiency, and, possibly, partial assimilation of the CNF by the animals. However, at low food levels and the highest CNF concentrations, growth and reproduction were negatively affected. These responses were linked to caloric restriction caused by dilution of the food source, but not an obstruction of the alimentary canal. Finally, no apparent translocation of CNF past the alimentary canal was detected. We conclude that CNF displays a low toxic potential to filter-feeding organisms and the expected environmental risks are low.     

Responses of Daphnia magna to pulsed exposures of arsenic

Research on the toxicity of arsenic has focused on sublethal effects that do not provide sufficient information for risk estimation. While most studies have focused on organism response to constant arsenic exposures, organisms in nature are exposed to fluctuating As concentrations. Consequently, results obtained from standardized bioassays with constant exposures may not adequately characterize risk to indigenous biota. This research was designed to characterize the response of Daphnia magna to fluctuating arsenic exposures during 21-day experiments. At concentrations > or =3000 microg/L As, 21-day pulsed exposure mortality increased as a function of exposure concentration and duration. In addition, 21-day pulsed exposure mortality increased with increasing recovery time. Pulsed As exposure did not affect the growth of D. magna over 21 days. Twenty-one day accumulative reproduction of D. magna was only affected by pulsed exposures of high As concentration and long durations.     

Chronic toxicity of a homologous series of linear alcohol ethoxylate surfactants to Daphnia magna in 21 day flow-through laboratory exposures

The chronic toxicity to Daphnia magna Straus in 21 day flow-through laboratory exposures was assessed for a homologous series of nonionic linear alcohol ethoxylate surfactants. Three primarily linear alcohol ethoxylate surfactants with alkyl chain distributions of C_9-11, C_12-13, and C_14-15, and with respective average ethoxymer groups of 6, 6.5, and 7 were tested. Mean measured surfactant exposure concentrations agreed with nominal concentrations, with average recoveries of 93, 84, and 86% (R²=0.99, 0.99, and 0.98) for the C_9-11EO6, C_12-13EO6.5, and C_14-15EO7 surfactants, respectively. Daphnia magna survival 21 day EC₅₀s were 5.9, 2.2 and 1.2 mg L⁻¹ for the C_9-11, C_12-13, and C_14-15 surfactants, respectively. Data indicate a twofold increase in toxicity in terms of daphnid survival with each two carbon addition in alkyl chain length. Daphnid survival and reproduction were equally sensitive for the C_9-11 and C_14-15 surfactants, however, reproduction was more sensitive than survival for the C_12-13 surfactant. Survival NOECs were 2.77, 1.75, and 0.79 mg L⁻¹ while reproduction no observed effect concentrations (NOECs) were 2.77, 0.77, and 0.79 mg L⁻¹ for the C_9-11, C_12-13, and C_14-15 surfactants, respectively. For the surfactants studied, chronic laboratory NOECs for D. magna survival (21 day) were similar to NOECs obtained for cladoceran densities in stream mesocosm experiments (30 day exposures) indicating a direct relationship between laboratory chronic effects and field effect levels for similar organisms. ©1999 John Wiley & Sons, Inc. Environ Toxicol 14: 293–300, 1999     

Reprint of "Chronic toxicity of silver nitrate to Ceriodaphnia dubia and Daphnia magna, and potential mitigating factors"

We investigated the chronic toxicity of Ag, as silver nitrate, using two freshwater aquatic cladoceran species, Ceriodaphnia dubia and Daphnia magna, to generate data for the development of a chronic ambient water quality criterion for Ag. Preliminary studies with C. dubia showed variable results which were related to the equilibration time between food and silver. Follow-up testing was conducted using a 3 h equilibration time, which stabilized dissolved Ag concentrations and the toxicity of Ag(+). Results with C. dubia conducted individually (1 per cup, n=10) and in mass (30 per chamber, n=2) gave similar results once similar standardized equilibration times were used. The maximum acceptable toxicant concentration (MATC) of Ag to C. dubia and D. magna was 9.61 and 3.00 microg dissolved Ag/L, respectively. The chronic toxicity of Ag(+) to C. dubia was also evaluated in the presence of: (1) dissolved organic carbon (DOC) and (2) sulfide. The addition of DOC (0.4 mg/L) resulted in a approximately 50% decrease in toxicity while the addition of sulfide (75.4 nM) deceased toxicity by 42%. Whole-body Ag concentration in D. magna was positively correlated with increased levels of Ag exposure, however; we observed a non-statistical decrease in whole-body Na levels, an estimator of sodium homeostasis.     

Ecotoxicological effects of bisphenol A and nonylphenol on the freshwater cladocerans Ceriodaphnia silvestrii and Daphnia similis

Toxicities of bisphenol A (BPA) and nonylphenol (NP) to the neotropical freshwater cladocerans Ceriodaphnia silvestrii and Daphnia similis were studied under laboratory conditions. Acute exposures to BPA generated mean 48-h EC₅₀ values of 14.44 (6.02–22.85) mg L^?1 for C. silvestrii and 12.05 (1.73–22.37) mg L^?1 for D. similis. When the organisms were exposed to acute doses of NP, mean 48-h EC₅₀ values were 0.055 (0.047–0.064) mg L^?1 (C. silvestrii) and 0.133 (0.067–0.200) mg L^?1 (D. similis). Ceriodaphnia silvestrii was also tested in chronic bioassays, which resulted in mean 8-d IC₂₅ values of 2.43 (2.16–2.69) mg L^?1 BPA [no observed effect concentration (NOEC): 1.38?mg L^?1] and 0.020 (0.015–0.026) mg L^?1 NP (NOEC: 0.015?mg L^?1). These laboratory tests are valuable to broaden the understanding of the environmental threat posed by BPA and NP in aquatic ecosystems, and to increase the knowledge about the sensitivity of neotropical indigenous species to these contaminants. In addition to the laboratory bioassays, species sensitivity distributions were used to suggest protective concentrations of BPA and NP to prevent adverse effects on freshwater organisms. According to the obtained results, concentrations lower than 36.47?µg L^?1 BPA and 1.39?µg L^?1 NP are not expected to adversely impact aquatic organisms in natural ecosystems.     

Toxicity of the cyanobacterium Cylindrospermopsis raciborskii to Daphnia magna

The effect of two strains of Cylindrospermopsis raciborskii on the survivorship, somatic growth, and detoxification processes of juvenile Daphnia magna were investigated. Both strains of C. raciborskii (and also Ankistrodesmus falcatus, used as the control) were given to newborn D. magna at equivalent biovolumes. The survival curves for D. magna subjected to the two C. raciborskii treatments differed from those of the starved and fed treatments. After 48 h of exposure, the percentage of D. magna surviving after exposure to Cylin-A (a cylindrospermopsin-producing strain isolated from Australia) and Cylin-P (a non-cylindrospermopsin-producing strain isolated from Portugal) was 10.00% and 93.33%, respectively. The strain that produces cylindrospermopsin caused the greatest toxic effect in juvenile D. magna. Statistically significant differences in D. magna body size between the four treatments (Cylin-A, Cylin-P, A. falcatus, and starved) were detected after 48 h of exposure. The juvenile D. magna that received the two C. raciborskii treatments showed an increase in size (relative to their size at T(0)) of 2.54% and 38.14%, respectively. These values were statistically significantly different than those of the A. falcatus-fed control (55.54%) and the starved control (11.47%). In both C. raciborskii treatments there was a tendency for increased GST enzyme activities after 24 h of exposure. Cylindrospermopsin was detected (HPLC-MS/MS) in D. magna tissues after 24 and 48 h (0.025 and 0.02 ng animal(-)1, respectively). The results of this study indicate that C. raciborskii can affect the fitness and growth potential of juvenile D. magna.     

pH‐Dependence and QSAR analysis of the toxicity of phenols and anilines to Daphnia magna

The toxicity of 13 anilines and 11 phenols to Daphnia magna has been assessed at pH values of 6.0, 7.8, and 9.0. Toxicity of anilines decreases with decreasing pH, whereas the toxicity of phenols decreases with increasing pH. These variations in toxicity, are clearly brought about by the effect of ionization, especially for the phenols. Toxicity decreased with an increasing proportion of ionized molecules. The relative contribution of the ionized and unionized molecules was assessed. It is believed that, at least for the phenols, the ionized form of the compounds does contribute to the toxicity of the molecules. However, the decreased overall toxicity suggests that a lower relative toxicity is attributable to the ionized form. Attempts were also made to develop quantitative structure‐activity relationship (QSAR) models for these toxicity data. Reasonable QSARs were produced for the anilines and phenols, acting as polar narcotics, considered together; these QSARs included a measure of hydrophobicity, pK_a, and hydrogen bonding. When the phenols and anilines were considered separately, better, hydrophobicity‐based, QSARs were obtained. Most compounds, with the exception of 2,4‐dinitrophenol, resorcinol, and pentachlorophenol, were thought to be acting by a polar narcosis mechanism of toxic action. There was some evidence that 2,4‐dichlorophenol and 2,4,6‐trichlorophenol may be acting as weak acid respiratory uncouplers, despite earlier suggestions that they act as polar narcotics. © 2000 John Wiley & Sons, Inc. Environ Toxicol 15: 140–148, 2000     

The developmental toxicity of 1-methyl-3-octylimidazolium bromide on Daphnia magna

The developmental toxicity of 1-methyl-3-octylimidazolium bromide ([C(8)mim]Br) on Daphnia magna was investigated. The 24 and 48 h LC(50) values for [C(8)mim]Br in D. magna were 1.99 and 0.95 mg/L, respectively. A series of multigenerational toxicity tests were then used to explore [C(8)mim]Br effects in D. magna. [C(8)mim]Br significantly inhibited the body lengths of the F0 and F1 1st generations. After 21 days of exposure, [C(8)mim]Br lowered the reproductive ability of the F0 and F1 1st generations. In F1 3rd generation, 21 days of [C(8)mim]Br exposure prolonged the time to bear the first egg and the time to the first brood compared with the control, but the number of first-brood offspring and the number of broods produced by these animals were reduced. After the recovery period all the reproductive parameters returned to normal in F1 1st generation except for the number of broods. The dead neonates increased with prolonged exposure and increasing concentrations, and the dead neonates of the F1 3rd generation went far beyond that of the F1 1st and F0 generations. The intrinsic rate of natural increase (r) values of the three D. magna generations significantly decreased after exposure to higher concentrations of [C(8)mim]Br compared with control groups. Collectively, these results suggest that [C(8)mim]Br exerts a toxic effect on the development of D. magna. This study also highlights the importance of systematically evaluating the potential effects of aquatic ecosystems of ionic liquids that may be released into bodies of water.     

The chronic toxicity of ZnO nanoparticles and ZnCl2 to Daphnia magna and the use of different methods to assess nanoparticle aggregation and dissolution

In this study, the effect of ZnO nanoparticles and ZnCl₂ on growth, reproduction and accumulation of zinc in Daphnia magna was determined in a 21-day chronic toxicity test. A variety of techniques were used to distinguish the free zinc ion, dissolved, nanoparticle and aggregated zinc fraction in the Daphnia test medium. The results showed similar chronic effects on growth, reproduction and accumulation for the ZnO nanoparticles (EC₁₀, ₂₀, ₅₀ reproduction: 0.030, 0.049, 0.112 mg Zn/l) and the ZnCl₂ (EC₁₀, ₂₀, ₅₀ reproduction: 0.014, 0.027, 0.082 mg Zn/l). A large fraction of the nanoparticles rapidly dissolved after introduction in the exposure medium. Aggregation of nanoparticles was also observed but within 48 h of exposure most of these ZnO aggregates were dissolved. Based on the combined dissolution kinetics and toxicity results, it can be concluded that the toxicological effects of ZnO nanoparticles at the chronic level can be largely attributed to the dissolved fraction rather than the nanoparticles or initially formed aggregates.     

Effects of copper pre-exposure routes on the energy reserves and subsequent copper toxicity in Daphnia magna

The hypothesis was tested that copper uptake routes affect the tolerance of Daphnia magna to copper and influence the energy reserves. These were determined in D. magna juveniles that had been exposed for 4 days to water borne and/or dietary copper (algae Pseudokichneriella subcapitata loaded with copper) at nominal concentrations of 0, 10, and 100 nM. Tolerance increased with dietary copper pre-exposure reflected in 24 and 48 h LC50 values of 466 and 398 nM at 100 nM pre-exposure versus 301 and 254 nM in controls, respectively. Control animals (no copper added to their exposure medium and diet) had the lowest lipid content and consequently the lowest energy content. The current study stresses the importance of addressing dietary exposure routes in metal toxicity assessments.     

Disaggregation of gold nanoparticles by Daphnia magna

Abstract

The use of manufactured nanomaterials is rapidly increasing, while our understanding of the consequences of releasing these materials into the environment is still limited and many questions remain, for example, how do nanoparticles affect living organisms in the wild? How do organisms adapt and protect themselves from exposure to foreign materials? How does the environment affect the performance of nanoparticles, including their surface properties? In an effort to address these crucial questions, our main aim has been to probe the effects of aquatic organisms on nanoparticle aggregation. We have, therefore, carried out a systematic study with the purpose to disentangle the effects of the freshwater zooplankter, Daphnia magna, on the surface properties, stability, and aggregation properties of gold (Au) nanoparticles under different aqueous conditions as well as identified the proteins bound to the nanoparticle surface. We show that Au nanoparticles aggregate in pure tap water, but to a lesser extent in water that either contains Daphnia or has been pre-conditioned with Daphnia. Moreover, we show that proteins generated by Daphnia bind to the Au nanoparticles and create a modified surface that renders them less prone to aggregation. We conclude that the surrounding milieu, as well as the surface properties of the original Au particles, are important factors in determining how the nanoparticles are affected by biological metabolism. In a broader context, our results show how nanoparticles released into a natural ecosystem become chemically and physically altered through the dynamic interactions between particles and organisms, either through biological metabolism or through the interactions with biomolecules excreted by organisms into the environment.     

Fitness parameters and DNA effects are sensitive indicators of copper-induced toxicity in Daphnia magna.

This study compared the effects occurring at molecular and population levels in Daphnia magna exposed to copper concentrations in the range of 15-120 microg/l. The qualitative and quantitative modifications arising in random amplified polymorphic DNA (RAPD) profiles as a measure of DNA effects were compared with a number of key ecological fitness parameters, namely, the age-specific survival, age-specific fecundity, net reproductive rate, and intrinsic rate of population increase. Results suggested that growth, reproduction, and most of the fitness parameters as well as genomic template stability (a qualitative measure reflecting changes in RAPD profiles) were significantly affected at copper concentrations of 90 and 120 microg/l. Among the fitness parameters, the age-specific fecundity and net reproductive rate were the most sensitive parameters of toxicity. Changes in RAPD patterns generally occurred at copper concentrations of 90 and 120 microg/l, but with one primer, changes significantly arose at all copper concentrations. Overall, molecular and population parameters compared well and represented a sensitive means to measure toxicity induced by copper in Daphnia magna. In conclusion, the measurement of parameters at both molecular and population levels is valuable for investigating the specific effects of agents interacting with DNA. Ultimately, this methodology may allow the ecotoxicological examination of the link between molecular alterations and measurable adverse effects at higher levels of biological organization.     

Toxicity evaluation of metal plating wastewater employing the Microtox assay: a comparison with cladocerans and fish

The relative sensitivity of the Microtox assay is closely related to the type of toxicant, and hence its utility in biomonitoring effluents is better evaluated on a case-by-case basis. The Microtox assay, employing the marine bacterium Vibrio fischeri, was evaluated for its applicability in monitoring metal plating wastewater for toxicity. The results of the Microtox assay after 5, 15, and 30 min of exposure, were compared with data obtained from conventional whole effluent toxicity testing (WET) methods that employed Daphnia magna, Ceriodaphnia dubia, and the fathead minnow (Pimephales promelas). The Microtox assay produced notably comparable EC50 values to the LC50 values of the acute fathead minnow toxicity test (< 0.5 order of difference). The Spearman's rank correlation analyses showed that the bacterial assay, regardless of exposure duration, correlated better with the acute fish than the daphnid results (p < 0.05). These observations were consistent to other studies conducted with inorganic contaminants. The relative sensitivity of the 30-min Microtox assay was within the range of the two frequently used acute daphnid/fish toxicity tests. In conclusion, the Microtox assay correlated well with the acute fathead minnow data and is well suited for toxicity monitoring for these types of industrial wastes.     

Effect of culture volume and adult density on the neonate production of Daphnia magna,as a test organism for aquatic toxicity tests

Neonates of Daphnia magna are widely used for bioassays in aquatic toxicology. Although this species is considered as a world‐wide reference, its low repeatability of the results in the ecotoxicological studies, are due to the differences in maintenance conditions of the brood stock. We studied the combined effects of adult parthenogenetic female density and culture volume on the total fecundity. Extrapolated densities from 5 to 40 adults L⁻¹, in volumes from 100 to 400 mL were tested. The green microalga Scenedesmus incrassatulus was used as food (1.30E+06 cells mL⁻¹). Once the reproduction began, the neonates produced were counted daily and discarded to avoid crowding effects. The experiments were carried out for 40 days. The maximal total offspring per female, i.e., 832 and 755 neonates, were obtained in the extrapolated densities of 5 and 10 ind. L⁻¹, respectively, using one adult female per culture vessel (in 200 and 100 mL of medium). When the quantity of the culture medium increased, despite a lower female density, the fecundity decreased and also the total offspring per female. For optimal neonate production, the parthenogenetic females should be individually cultured in small volumes (100 or 200 mL). © 2000 John Wiley & Sons, Inc. Environ Toxicol 15: 155–159, 2000