Effect assessment of nano- and microplastics in freshwater ecosystems

The release of plastic waste into the environment has become one of the major water quality problems of the Anthropocene. Nanoplastics, with a size smaller than 0.1 µm, and microplastics, with a size between 0.1 µm and 5 mm, comprise the smallest particle fraction of plastic debris. Although the accumulation of nano- and microplastics is currently a major concern, studies addressing their effects on single species are scarce and nothing is known about their long-term effects at the community level. Freshwaters are particularly affected as sediments are known to accumulate nano- and microplastics due to the vicinity of sources and due to aggregation and biofouling processes and subsequent settling, which create hotspot areas that might pose a risk for benthic organisms. The general objective of this thesis is to assess threshold effect concentrations of nano- and microplastics in freshwater ecosystems, and to provide guidance on how such effects should be assessed in the context of risk assessment. For this, chronic single species tests were conducted with freshwater benthic macroinvertebrates exposed to nano- and microplastics via sediment following a standard setup to measure their individual threshold-effect concentrations. Benthic macroinvertebrates were exposed to polystyrene microplastic fragments, microplastics made by scraping and grinding second-hand tires and raspberry-shaped polystyrene nanoplastics with a palladium-core, separately. The tested benthic macroinvertebrates included the amphipods Gammarus pulex and Hyalella azteca, the isopod Asellus aquaticus, the worms Lumbriculus variegatus and Tubifex spp., and the bivalve Sphaerium corneum. The ingestion, retention and egestion of the polystyrene microplastics, the microplastics released from car tyres and the polystyrene nanoplastics were analysed for G. pulex using micro-Fourier Transformed Spectroscopy, image analysis and inductively coupled plasma mass spectrometry, respectively. In addition to the effects of nano- and microplastics on single species, we measured community threshold-effect concentrations for nano- and microplastics. For this, trays containing sediment and nano or microplastics at five concentrations, including two environmentally realistic concentrations, were embedded at the bottom of a semi-artificial ditch containing a stable donor community. The donor community was allowed to colonize the trays and after 3 and 15 months, trays were retrieved, and species were identified and counted. Effects were assessed on the community composition, population sizes and species diversity. In this thesis, we also explore the potential use of in vitro toxicity tests to assess the risks of nanoplastics on human health, in this case with and without chemical mixtures originating from wastewater treatment plant effluent and surface waters. To this aim, we evaluated the genotoxicity of two spherical polystyrene nanoplastics with average sizes of 50 nm and 500 nm at four concentrations in three matrices using the Ames fluctuation test, which has the purpose of detecting base-pair and frameshift mutations in the genome of Salmonella typhimurium with and without metabolic activation. Then, following recently developed quality assessment methods for studies reporting abundance of microplastic in biota and water samples, we critically review 105 papers reporting microplastic effects on aquatic biota. For this, 20 Quality assurance/Quality control (QA/QC) criteria were defined within four main categories: particle characterization, experimental design, applicability for risk assessment and ecological relevance. With the aim of detecting knowledge gaps within effect studies with microplastic, we provide an overview of the study characteristics of the reviewed studies with respect to the size, shape and polymer type of the microplastic used, the tested species, the duration of the exposure, the endpoints studied and use or not of effect thresholds to report the results. Moreover, demonstrated and suggested effects and effect mechanisms reported in the reviewed papers are summarized and discussed, with the results of the quality evaluation applied as a method to assess the overall weight of evidence regarding probable ecologically relevant effects. Finally, we develop a tiered approach ecological risk assessment for microplastic using the data generated in this thesis and data taken from the literature.