Increasing amounts of engineered nanoparticles (NPs) in wastewater can reach the aquatic environment by passing through the sewage treatment plant (STP). NPs can induce ecotoxicological effects due to their specific chemical properties. However, their bioavailability and toxicity are potentially influenced by transformation processes caused by substances present in the STP, e.g., humic acids or sulfides. Due to the lack of a test system allowing to test NPs under realistic environmental conditions, we coupled two existing test systems, the activated sludge simulation test (OECD TG 303A 2001) and the chronic exposure test with the freshwater amphipod Hyalella azteca (Environment Canada 2013), to gain a test scenario that allows to consider the altered behavior and fate of NPs induced by the STP process. This should improve the environmental realism of the chronic exposure test with Hyalella. In the first study, we tested the STP effluent containing AgNPs. In the second and third study, tap water and control STP effluent were spiked with AgNPs and used as test media. The chronic exposure studies with the freshwater amphipod H. azteca showed that the investigated AgNPs lose most of their toxicity while passing through the STP. Over all studies with total Ag concentrations ranging from 0.85 to 68.70 µg/L, significant effects of the AgNPs were only observed in the survival of test animals exposed to tap water containing the highest Ag concentration (62.59 µg/L). Accumulation of silver in the body of test animals was clearly dependent on the pretreatment of the AgNPs. Silver ions (Ag+) released from AgNPs are supposed to be the major pathway leading to body burden following exposure to test media containing AgNPs. The coupled test system is suitable for testing substances that can reach the environment via the STP effluent. The investigated AgNPs lose most of their toxicity while passing through the STP. Accumulation of silver in the animals exposed to the different treatments was apparent, whereby silver ions (Ag+) released from AgNPs were supposed to be the major pathway leading to body burden.

Even though nanoparticles (NPs) are mostly removed by wastewater treatment plants, wastewater-borne NPs may show an altered toxicity to aquatic organisms. The main objectives of this work were: i) to assess the chronic (28 days) effects of wastewater-borne NPs of silver (AgNPs, 1.4-36.2 μg L-1) and titanium dioxide (TiO2NPs, 3.1-50.2 μg L-1) at the individual (growth) and biochemical (biomarkers of neurotoxicity, oxidative stress and energy metabolism) levels in rainbow trout Oncorhynchus mykiss; and ii) to compare them with their effluent-supplemented and water-dispersed counterparts. The total Ag and Ti levels were determined in several fish organs. The growth of O. mykiss was not affected by the NPs in any treatment, except a 29% increase at 5.5 μg L-1 of total Ag supplemented to effluents. The Ag level in organs of O. mykiss was significantly higher after exposure to water-dispersed AgNPs than their wastewater-borne or effluent-supplemented counterparts. No significant Ti uptake could be observed. Effluent-supplemented TiO2NPs (50.1 μg L-1 Ti) potentially induced neurotoxic effects, indicated by a 24% increase in acetylcholinesterase activity comparatively to controls. Energy reserves were unaffected by TiO2 treatments, while nearly all AgNP-containing treatments caused a depletion of total lipids, proteins and carbohydrates in the muscle, suggesting an increased energy demand for detoxification processes to cope with AgNPs. Besides NPs, the effluent matrix and dispersing agent (for AgNPs) induced significant effects on energetic reserves and oxidative stress, indicating background toxicity of both treatments at the biochemical level. Our study is the first to assess chronic effects of wastewater-borne NPs on rainbow trout. While no effects were found at the individual level, several biochemical markers were changed by the NPs exposure. Our results highlight the importance of using complex matrices for a reliable risk assessment of NPs in the aquatic environment.