Sensitivities of Australian and New Zealand amphipods to copper and zinc in waters and metal-spiked sediments

The sensitivities of eight benthic amphipods, Chaetocorophium cf. lucasi, Corophium colo, Grandidierella japonica, Hyale crassicornis, Hyale longicornis, Melita awa, Melita matilda and Melita plumulosa, to copper and zinc in water-only and whole-sediment toxicity tests were compared. Whole-sediment tests used copper- (1300 mg/kg) and zinc- (4000 mg/kg) spiked sediments after equilibration for sufficient time to produce pore water and overlying water concentrations below the lowest observable effect concentrations of water-only exposures. Survival of adults (after 10 d) and juveniles (after 96 h), and the metal concentrations in the body tissues of adults, were determined at the end of the tests. Two epibenthic amphipods from the genus Melita were the most sensitive species to aqueous copper and zinc, with a 96-h LC50 value of 120 microg Cu/l for both M. awa and M. plumulosa juveniles, and a 96-h LC50 value of 640 microg Zn/l for juveniles of M. plumulosa. Juvenile amphipods (7-d old) were more sensitive than adult amphipods (>30-d old) in both water-only and whole-sediment tests, with adult-LC50/juvenile-LC50 ratios in water-only tests ranging from 1.2 to l.5 for copper and 1 to 1.4 for zinc. All species except C. colo, C. cf. lucasi and M. matilda were sensitive to the copper-spiked sediment, with survival between 14% and 74% of controls. Similarly, all species except C. colo and G. japonica, showed a response to the zinc-spiked sediment (26-81% of control survival). The epibenthic amphipods were more sensitive than the infaunal tube-dwelling amphipods and are recommended as test species.     

Inter- and intra-species variation in acute zinc tolerance of field-collected cladoceran populations

Acute zinc toxicity was assessed for 10 freshwater cladoceran species collected in six different ecosystems across Europe and for two standard laboratory-reared species (Daphnia magna and Ceriodaphnia dubia). The collected organisms belonged to five different genera: Daphnia (subgenus Daphnia and Ctenodaphnia), Ceriodaphnia, Simocephalus, Acroperus and Chydorus. The 48-h EC50 of the field-collected organisms tested in standard laboratory water ranged from 375+/-141 to 4314+/-1513 microg Znl(-1). The laboratory clone of D. magna was less sensitive than the majority of the field-collected species, while our laboratory Ceriodaphnia dubia was the second most sensitive. Considerable inter-species variation was found within the genus of Ceriodaphnia (factor 6) and within the genus Daphnia (factor 8). Among the different (sub)genera tested, Chydorus and Ctenodaphnia were significantly more tolerant than the others (up to a factor 3 difference). A significant positive relationship (r2=0.67, p<0.05) between the mean cladoceran 48-h EC50 and the ambient zinc concentration of the different aquatic systems was demonstrated, suggesting a role of acclimation and/or adaptation. No significant correlation between the acute zinc tolerance and the length of the organisms was found.     

Lead,zinc and copper accumulation and tolerance in populations of Paspalum distichum and Cynodon dactylon

Both Fankou and Lechang lead/zinc (Pb/Zn) mine tailings located at Guangdong Province contained high levels of total and DTPA-extractable Pb, Zn and Cu. Paspalum distichum and Cynodon dactylon were dominant species colonized naturally on the tailings. Lead, zinc and copper accumulation and tolerance of different populations of the two grasses growing on the tailings were investigated. Tillers of these populations including those from an uncontaminated area were subjected to the following concentrations: 5, 10, 20, 30 and 40 mg l(-1) Pb, 2.5, 5, 10, 20 and 30 mg (l-1) Zn, or 0.25, 0.50, 1 and 2 mg l(-1) Cu for 14 days, respectively, then tolerance index (TI) and EC50 (the concentrations of metals in solutions which reduce 50% of normal root growth) were calculated. The results indicated that both Lechang and Fankou populations of the two grasses showed a greater tolerance to the three metals than those growing on the uncontaminated area, which suggested that co-tolerant ecotypes have evolved in the two grasses. P. distichum collected from Fankou tailings had the highest tolerance to Cu while Lechang population the highest tolerance to Pb and Zn among the tested populations, and tolerance levels in P. distichum were related to metal concentrations in the plants. P. distichum had a better growth performance than C. dactylon when both of them were grown on the tailings sites. Tolerant populations of these species would serve as potential candidates for re-vegetation of wastelands contaminated with Pb, Zn and Cu.     

Models for the field-based toxicity of copper and zinc salts to wheat in 11 Australian soils and comparison to laboratory-based models

Laboratory-based relationships that model the phytotoxicity of metals using soil properties have been developed. This paper presents the first field-based phytotoxicity relationships. Wheat (Triticum aestivum L.) was grown at 11 Australian field sites at which soil was spiked with copper (Cu) and zinc (Zn) salts. Toxicity was measured as inhibition of plant growth at 8 weeks and grain yield at harvest. The added Cu and Zn EC10 values for both endpoints ranged from approximately 3 to 4760 mg/kg. There were no relationships between field-based 8-week biomass and grain yield toxicity values for either metal. Cu toxicity was best modelled using pH and organic carbon content while Zn toxicity was best modelled using pH and the cation exchange capacity. The best relationships estimated toxicity within a factor of two of measured values. Laboratory-based phytotoxicity relationships could not accurately predict field-based phytotoxicity responses.     

Combustion experiments using pentachlorophenol on a pilot scale and full-scale

Chlorinated phenols are used at many saw mills for impregnation of saw timber. Waste products containing the used compound follow this process. Tests have been performed on pilot and full-scale in order to investigate the possibility of destroying these wastes by combustion.The pilot scale tests with 70 g of pentachlorophenol show that at a transit time of approximately 3 seconds and a combustion temperature of 600°C the unburnt residue was 50 mg/kg fed in pentachlorophenol. At 800°C the residue decreased to 15 mg/kg. No formation of octachlorodioxin could be proved.The full-scale test with a small amount of pentachlorophenol and a large amount of tetrachlorophenol show that at a transit time of approximately 0.9 seconds and a temperature varying during one test between 620–875° (average 760°C), the unburnt residue was 9 mg/kg for tetrachlorophenol and 650 mg/kg for pentachlorophenol. At an increased load the residue increased gradually, being 5.900 mg/kg for tetrachlorophenol and 7.100 mg/kg for pentachlorophenol at a test when the temperature was varied between 460 and 850° (average 620°C). Analyses of chlorinated dioxins show that octachlorodioxin was found in the flue gas at one of the tests. This may indicate a formation.     

Effects of six selected antibiotics on plant growth and soil microbial and enzymatic activities

The potential impact of six antibiotics (chlortetracycline, tetracycline and tylosin; sulfamethoxazole, sulfamethazine and trimethoprim) on plant growth and soil quality was studied by using seed germination test on filter paper and plant growth test in soil, soil respiration and phosphatase activity tests. The phytotoxic effects varied between the antibiotics and between plant species (sweet oat, rice and cucumber). Rice was most sensitive to sulfamethoxazole with the EC10 value of 0.1 mg/L. The antibiotics tested inhibited soil phosphatase activity during the 22 days' incubation. Significant effects on soil respiration were found for the two sulfonamides (sulfamethoxazole and sulfamethazine) and trimethoprim, whereas little effects were observed for the two tetracyclines and tylosin. The effective concentrations (EC10 values) for soil respiration in the first 2 days were 7 mg/kg for sulfamethoxazole, 13 mg/kg for sulfamethazine and 20 mg/kg for trimethoprim. Antibiotic residues in manure and soils may affect soil microbial and enzyme activities.     

Toxicity of the anthelmintic abamectin to four species of soil invertebrates

Abamectin is a veterinary medicinal product with high efficiency against parasitics. This study evaluates the sublethal toxicity of abamectin to three groups of soil dwelling organisms (springtails, enchytraeids and earthworms). The reproduction of the two springtail species Folsomia fimetaria and Folsomia candida was significantly affected at abamectin concentrations of 0.25 and 0.5 mg/kg DW, respectively. Adults of especially F. candida were less sensitive. Abamectin had no effect on the survival of enchytraeids, whereas the number of juveniles was reduced at concentrations above 10 mg/kg. The EC10 values for reproduction were in the range of 0.05 mg/kg for springtails and 12.8 mg/kg for enchytraeids. Abamectin also had a significant effect on the reproduction of earthworms with an EC10 value of 0.06 mg/kg. However, no change in the survival and growth of juvenile earthworms was observed following 70 days of exposure.     

Comparative sensitivity of 20 bioassays for soil quality

Increasing evidence suggests that the use of a single bioassay will never provide a full picture of the quality of the environment. Only a test battery, composed of bioassays of different animal and plant species from different trophic levels will reduce uncertainty, allowing an accurate assessment of the quality of the environment. In the present study, a test battery composed of 20 bioassays of varying biological endpoints has been compared Apart from lethality and reproductive failure in earthworms, springtails, nematoda, algae and vascular plants, these endpoints also included bioavailibility of metals (bacteria), heat-shock induction (nematodes, algae), DNA damage (bacteria, earthworm, vascular plants), β-galactosidase (Daphnia) and esterase activity (algae) and a range of immunological parameters (earthworm). Four chemicals (cadmium, phenol, pentachlorophenol and triflurahn) — each representing a different toxic mode of action — were applied in a dilution series (from 1 mg/kg up to 1000 mg/kg) onto OECD standard soil. The tests have been performed both on these artificially contaminated soil samples and on aqueous extracts subsequently obtained from these soils. The results show that the immunological parameters and the loss of weight in the earthworms were among the most sensitive solid phase assays. Esterase inhibition and heat-shock induction in algae were shown to be extremely sensitive when applied to soil extracts. As previously shown at the species level, no single biological endpoint was shown to be the most sensitive for all four modes of toxic action.     

Performance of different microalgal species in removing nickel and zinc from industrial wastewater

A series of batch experiments was conducted to compare the ability of 11 microalgal species of the same cell density in removing nickel (Ni) and zinc (Zn) from synthetic wastewater. These included Chlorella vulgaris (commercially available), Chlorella sorokiniana and Scenedesmus quadricauda (isolates from polluted water of Wuhan, China), and eight different isolates from Hong Kong. The Wuhan isolate of Scenedesmus removed most Ni, probably due to its large biomass. Nickel concentration was reduced from an initial 30 to 0.9 mg/l after 5 min (97% Ni removal), and further declined to 0.4 mg/l after 90 min of treatment. In wastewater containing 30 mg/l Ni and 30 mg/l Zn, more than 98%, Ni and Zn were removed simultaneously at the end of 5 min treatment, indicating that the presence of Zn in wastewater did not affect Ni removal by this Scenedesmus isolate. The second most effective species for Ni removal was an isolate, tentatively identified as Chlorella miniata, Ni concentration was reduced to 10 mg/l after 90 min, and was only slightly interfered by the presence of Zn. In terms of metal removal per unit biomass or unit surface area of algal cells, C. miniata was the best species in removing Ni and Zn. At the other extreme, one Hong Kong isolate (Synechocystis sp.) did not remove any Ni and only achieved 40% Zn removal. Performance of the other isolates was comparable with the commercial C. vulgaris, less than 50% Ni was removed after 5 h of treatment and Ni removal was significantly reduced by the presence of Zn. All algae tested were found to be viable, showing these 11 species could tolerate a mixture of 30 mg/l Ni and 30 mg/l Zn in wastewater.     

Food chain transfer of cadmium and lead to cattle in a lead–zinc smelter in Guizhou, China

Cadmium (Cd) and Lead (Pb) are environmental pollutants. Environmental samples and bovine tissues were collected from the areas around a lead–zinc smelter in Guizhou, China for Cd, Pb, zinc (Zn) and copper (Cu) analysis. Cd in soil (10 mg/kg) and feed (6.6 mg/kg) from the polluted areas was 10 times higher than the Chinese Standards, resulting in higher Cd in bovine kidney (38 mg/kg) and liver (2.5 mg/kg). Pb in feed (132 mg/kg) from the polluted area was much higher than unpolluted areas, causing higher Pb levels in bovine tissues. Environmental Zn was elevated, but bovine tissue Zn was normal. Cu in bovine liver decreased with increased Cd and Pb. Metals in drinking water and in bovine muscle were within the Standard range. Thus, in the areas of this lead–zinc smelter, the environment has been contaminated with Cd and Pb, which has been transferred to cattle through the food chain.     

Metal-induced tolerance in the freshwater microbenthic diatom Gomphonema parvulum

The benthic diatom Gomphonema parvulum Kützing is a common species in both clean and metal contaminated rivers. Our aim was to investigate whether metal-induced tolerance could explain the persistance of this taxon under metal polluted conditions. G. parvulum strains were isolated from a Zn- and Cd-contaminated stream and from a relatively clean ("reference") stream. The strains were cultured in synthetic medium as mono-specific biofilms to maintain their specific benthic growth features. Moreover, the strain from the metal polluted stream was cultured in plain and Zn- and Cd-enriched synthetic medium. Short-term (5 h) toxicity experiments with Zn were performed with the strains using pulse amplitude modulated (PAM) fluorometry. Zn lowered significantly the minimal chlorophyll fluorescence (F0) and the photon yield (phi(p)) of the exposed strains after 5 h exposure. The actual Zn concentrations that caused a 50% reduction (EC50's) of the phi(p) of the strain from the metal polluted stream were significantly higher than those of the isolate from the unpolluted stream. The absence of tolerance to Cu of the "polluted" strain indicated that Zn tolerance resulted from specific induction by chronic exposure to Zn in the field. Observations on field biofilms confirmed a higher tolerance of the G. parvulum population from the polluted stream than of the G. parvulum population from the reference stream. A genetic nature of this metal adaptation was supported by the persistance of the Zn tolerance of the polluted strain 2 years after isolation.     

The toxicity of copper,cadmium and zinc to four different Hydra (Cnidaria: Hydrozoa)

An acute toxicity study of three metals to Hydra species carried out using two different assessment methods, (i) determination of the LC50 and (ii) measurement of progressive morphological changes, demonstrated that relative toxicity decreased from copper to cadmium with zinc the least toxic for all species. The latter method revealed more details of the effect on Hydra in terms of physical damage to the polyp but both methods indicated that H. viridissima was more sensitive to copper and cadmium than H. vulgaris¹ (Zurich strain, male clone), H. vulgaris² (a dioecious strain reproducing sexually and asexually) and H. oligactis (dioecious, reproducing sexually and asexually). The responses to zinc were similar for all Hydra. The possible role of metabolic interactions between H. viridissima and its symbiotic green algae in contributing to the greater sensitivity of this polyp is discussed.     

Toxicity of abamectin and doramectin to soil invertebrates

This study aimed at determining the toxicity of avermectins to soil invertebrates in soil and in faeces from recently treated sheep. Abamectin was more toxic than doramectin. In soil, earthworms (Eisenia andrei) were most affected with LC50s of 18 and 228 mg/kg dry soil, respectively, while LC50s were 67-111 and >300 mg/kg for springtails (Folsomia candida), isopods (Porcellio scaber) and enchytraeids (Enchytraeus crypticus). EC50s for the effect on reproduction of springtails and enchytraeids were 13 and 38 mg/kg, respectively for abamectin, and 42 and 170 mg/kg for doramectin. For earthworms, NOEC was 10 and 8.4 mg/kg for abamectin and doramectin effects on body weight. When exposed in faeces, springtails and enchytraeids gave LC50s and EC50s of 1.0-1.4 and 0.94-1.1 mg/kg dry faeces for abamectin and 2.2->2.4 mg/kg for doramectin. Earthworm reproduction was not affected. This study indicates a potential risk of avermectins for soil invertebrates colonizing faeces from recently treated sheep.     

Toxic effects of anthraquinone and phenanthrenequinone upon Scenedesmus strains (green algae) at low and elevated concentration of CO2

Short-term (24h) experiments were performed to examine the effect of anthraquinone (ANTQ) and phenanthrenequinone (PHEQ) on two Scenedesmus armatus strains (B1-76 and 276-4d) grown in a batch culture system aerated with CO2 at a low (0.1%) or elevated (2%) concentration. ANTQ at concentrations within the range of 0.156-1.250 mg dm-3 inhibited the growth of B1-76 population in a concentration-dependent manner, and calculated EC50 for low-CO2 cells was 0.56 mg dm-3. The toxic effect of ANTQ on this strain was more pronounced in high-CO2 cells, where not only growth but also photosynthesis, respiration and SOD activity were significantly inhibited. In contrast, except for SOD activity, no ANTQ effects on strain 276-4d were found. PHEQ at concentrations within the range of 0.063-0.125 mg dm-3 inhibited the growth of B1-76 population in a concentration-dependent manner. The value of EC50 for low-CO2 B1-76 cells was 0.10 mg dm-3. PHEQ inhibited the growth of both strains regardless of CO2 concentration. In B1-76 cells affected by PHEQ, inhibition of photosynthesis was independent of the CO2 level, whereas the SOD activity was much higher in cultures aerated with 2% than with 0.1% CO2. Higher toxicity of PHEQ to strain 276-4d grown at 2% CO2 was accompanied by strong inhibition of photosynthesis, while in low-CO2 cells this process was slightly stimulated. The SOD activity in both low- and high-CO2 cells of strain 276-4d treated with PHEQ was 2-3 times higher compared with the controls. The pattern of SOD isoforms (PAGE analysis) obtained from cells exposed to ANTQ or PHEQ did not change compared with the controls, but the location of the SOD isoforms bands on gel was affected by the concentration of CO2. The results suggest that the strain-specific toxicity of ANTQ and PHEQ may result from oxidative stress. In addition, carbon dioxide appears to play an important role in the toxicity of quinones to algae.     

Biodegradation of phenol and phenol-related compounds by psychrophilic and cold-tolerant alpine yeasts

We characterized 32 cold-adapted, psychrophilic and cold-tolerant, yeast strains isolated from alpine habitats with regard to their taxonomy, growth temperature profile, and ability to degrade phenol and 18 phenol-related mono-aromatic compounds at 10 degrees C. Twenty of the strains were identified by sequencing of the ribosomal ITS region as seven species of the basidiomycota: Cryptococcus terreus (three strains), Cryptococcus terricola (one strain), Rhodosporidium lusitaniae (two strains), Rhodotorula creatinivora (10 strains), Rhodotorula ingeniosa (one strain), Mastigobasidium intermedium (one strain), and Sporobolomyces roseus (two strains). Twelve strains sharing closely related ITS sequences could not be identified to the species level; according to their ITS sequence they are included in the Microbotryomycetidae. These 12 strains were psychrophilic (no growth at temperatures above 20 degrees C); one-third of these strains did not grow above 15 degrees C. None of the 32 strains utilized any of the highly volatile mono-aromatic compounds (benzene, toluene, ethylbenzene, nitrobenzene, o-xylene, m-xylene, and p-xylene) as the sole carbon source. Non/low volatile aromatic compounds were degraded in the following order: phenol>hydroquinone>resorcinol>benzoate>catechol>salicylate>p-cresol>m-cresol. o-Cresol, guaiacol, p-nitrophenol, or p-nitrotoluene were not utilized for growth. R. creatinivora strains degraded up to seven compounds, whereas C. terricola and S. roseus strains degraded only two compounds. The toxicity of the compounds was determined via growth inhibition in the presence of toxicants and nutrients at 10 degrees C. R. creatinivora strains were characterized by higher IC50 values than other species, S. roseus was the most sensitive species. The most toxic compounds were the xylene isomers, ethylbenzene, p-nitrophenol, and m-cresol. There was a relation between the chemical structure of the compounds and their toxicity, whereas a relation between the toxicity of the compounds and the ability of the yeasts strains to utilize these compounds for growth was only detected in some cases.     

Phytotoxicity of nanoparticles: inhibition of seed germination and root growth

Plants need to be included to develop a comprehensive toxicity profile for nanoparticles. Effects of five types of nanoparticles (multi-walled carbon nanotube, aluminum, alumina, zinc, and zinc oxide) on seed germination and root growth of six higher plant species (radish, rape, ryegrass, lettuce, corn, and cucumber) were investigated. Seed germination was not affected except for the inhibition of nanoscale zinc (nano-Zn) on ryegrass and zinc oxide (nano-ZnO) on corn at 2000 mg/L. Inhibition on root growth varied greatly among nanoparticles and plants. Suspensions of 2000 mg/L nano-Zn or nano-ZnO practically terminated root elongation of the tested plant species. Fifty percent inhibitory concentrations (IC50) of nano-Zn and nano-ZnO were estimated to be near 50mg/L for radish, and about 20mg/L for rape and ryegrass. The inhibition occurred during the seed incubation process rather than seed soaking stage. These results are significant in terms of use and disposal of engineered nanoparticles.     

Vegetation establishment on a deposit of zinc mine wastes

Field trials concerning the establishment of plant cover on a deposit of wastes from the Ammeberg zinc mine in central Sweden were carried out during 1976-1985. Different soil conditioners and manures were applied and plant species cultivars were evaluated with regard to plant biomass, vigour, durability and content of zinc, lead and cadmium. Sewage sludge and topsoil led to better establishment of grasses than did municipal waste, straw and hydraulic seeding. After 2 years, Festuca rubra and Poa pratensis dominated the swards. Other species (Dactylis glomerata, Bromus inermis, Lolium perenne, Phleum nodosum, Festuca pratensis and F. arundinacea) constituted only a minor part of the stand. After 10 years, F. rubra was the most dominant species, while native Agrostis tenuis had invaded 20-50% of the area within the plots. Merlin was the clearly dominant red fescue cultivar. The concentration of zinc in shoots (616 mg kg(-1) dw) was about 10% of that in the soil. Zinc concentration decreased with increasing biomass above ground. It increased with age in Scots pine needles and was very high in birch leaves. Grasses survived longer than legumes in the zinc sand waste. Among the surviving grasses was a group with high (3800 mg kg(-1) dw) and a group with low (320 mg kg(-1) dw) zinc concentrations. The low group included Merlin red fescue and Sobel creeping bent. The cultivar Merlin contained a much lower zinc concentration than the other cultivars of red fescue (375 and 624 mg kg(-1) dw, respectively). A large amount of root biomass was present in plots with dominating Merlin red fescue (1715 g m(-2)), 97% of which was concentrated in the top 10 cm of the soil. The concentration of zinc in the roots was very high (13 000-25 000 mg kg(-1) dw). Nitrate fertilizer, especially ammonium nitrate, and acidic water (pH 4.3) increased zinc leaching.     

Effect of metal tolerant plant growth promoting Bradyrhizobium sp. (vigna) on growth, symbiosis, seed yield and metal uptake by greengram plants

The nickel and zinc tolerant plant growth promoting Bradyrhizobium sp. (vigna) RM8 was isolated from nodules of greengram, grown in metal contaminated Indian soils. The plant growth promoting (PGP) potentials of strain RM8 was assessed both in the presence and absence of nickel and zinc under in vitro conditions. Strain RM8 tolerated a high level of nickel (300 microg ml(-1)) and zinc (1400 microg ml(-1)) on yeast extract mannitol agar medium. Bradyrhizobium sp. (vigna) strain RM8 produced 13.3 microg ml(-1) of indole acetic acid in Luria Bertani broth at 100 microg ml(-1) of tryptophan which increased to 13.6 microg ml(-1) at 50 microg Ni ml(-1) and 13.5 microg ml(-1) at 300 microg Zn ml(-1). Strain RM8 was positive for siderophore, HCN and ammonia both in the absence and presence of nickel and zinc. The PGP activity of this strain was further evaluated with increasing concentrations of nickel and zinc using greengram as a test crop. The bioinoculant enhanced the nodule numbers by 82%, leghaemoglobin by 120%, seed yield by 34%, grain protein by 13%, root N by 41% and shoot N by 37% at 290 mg Ni kg(-1) soil. At 4890 mg Zn kg(-1) soil, the bioinoculant increased the nodule numbers by 50%, leghaemoglobin by 100%, seed yield by 36%, grain protein by 13%, root N by 47% and shoot N by 42%. The bioinoculant strain RM8 reduced the uptake of nickel and zinc by plant organs compared to plants grown in the absence of bioinoculant. This study suggested that the bioinoculant due to its intrinsic abilities of growth promotion and attenuation of the toxic effects of nickel and zinc could be exploited for remediation of metal from nickel and zinc contaminated sites.     

Chronic toxicity of ZnO nanoparticles, non-nano ZnO and ZnCl2 to Folsomia candida (Collembola) in relation to bioavailability in soil

The chronic toxicity of zinc oxide nanoparticles (ZnO-NP) to Folsomia candida was determined in natural soil. To unravel the contribution of particle size and free zinc to NP toxicity, non-nano ZnO and ZnCl₂ were also tested. Zinc concentrations in pore water increased with increasing soil concentrations, with Freundlich sorption constants K_f of 61.7, 106 and 96.4 l/kg (n = 1.50, 1.34 and 0.42) for ZnO-NP, non-nano ZnO and ZnCl₂ respectively. Survival of F. candida was not affected by ZnO-NP and non-nano ZnO at concentrations up to 6400 mg Zn/kg d.w. Reproduction was dose-dependently reduced with 28-d EC50s of 1964, 1591 and 298 mg Zn/kg d.w. for ZnO-NP, non-nano ZnO and ZnCl₂, respectively. The difference in EC50s based on measured pore water concentrations was small (7.94-16.8 mg Zn/l). We conclude that zinc ions released from NP determine the observed toxic effects rather than ZnO particle size.