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100Effective identification of species using short DNA fragments (DNA barcoding and DNA 101 metabarcoding) requires reliable sequence reference libraries of known taxa. Both 102 taxonomically comprehensive coverage and content quality are important for sufficient 103 accuracy. For aquatic ecosystems in Europe, reliable barcode reference libraries are 104 particularly important if molecular identification tools are to be implemented in biomonitoring 105 and reports in the context of the EU Water Framework Directive (WFD) and the Marine 106Strategy Framework Directive (MSFD). We analysed gaps in the two most important 107 reference databases, Barcode of Life Data Systems (BOLD) and NCBI GenBank, with a 108 focus on the taxa most frequently used in WFD and MSFD. Our analyses show that 109 coverage varies strongly among taxonomic groups, and among geographic regions. In 110 general, groups that were actively targeted in barcode projects (e.g. fish, true bugs, 111 caddisflies and vascular plants) are well represented in the barcode libraries, while others 112 have fewer records (e.g. marine molluscs, ascidians, and freshwater diatoms). We also 113 found that species monitored in several countries often are represented by barcodes in 114 reference libraries, while species monitored in a single country frequently lack sequence 115 records. A large proportion of species (up to 50%) in several taxonomic groups are only 116represented by private data in BOLD. Our results have implications for the future strategy to 117 fill existing gaps in barcode libraries, especially if DNA metabarcoding is to be used in the 118 monitoring of European aquatic biota under the WFD and MSFD. For example, missing 119 species relevant to monitoring in multiple countries should be prioritized. We also discuss 120 why a strategy for quality control and quality assurance of barcode reference libraries is 121 needed and recommend future steps to ensure full utilization of metabarcoding in aquatic 122 biomonitoring. 123 124

As documented by the numerous publications that have appeared in recent years, plastic pollution of the environment and the effects on the respective ecosystems are currently one of the most intensely discussed issues in environmental science and in society at large. Of special concern are the effects of micro-and nano-sized plastics. A key issue in understanding the fate and potential effects of micro-and nano-sized plastics is their dynamic nature, as the size, shape, and charge of the particles change over time. Moreover, due to various biological processes, such as the aggregation of organic material and/or bacteria ("biofouling"), the density of plastic particles that settle in the sediments of aquatic ecosystems may be several orders of magnitudes higher than that in the surrounding waters. Consequently, the risk posed by plastic pollution to benthic fauna is considerably high. Nonetheless, the vast majority of studies examining the effects of microplastics have focused on pelagic organisms so far. We therefore conducted a comprehensive literature review to examine the impact of micro-and nano-sized plastics on benthic invertebrates, including the physical and chemical effects of leaching and the interactions of plastic particles with contaminants. Overall, 330 papers were reviewed for their fulfillment of different criteria (e.g., test species, plastic material, particle shape, particle size, exposure concentration, exposure route, assay type, assay duration), with 49 publications finally included in our survey. A comprehensive gap-analysis on the effects of plastic particles on benthic invertebrates revealed a wide variety of effects triggered by micro-and/or nano-sized plastics but also distinct differences regarding the plastic materials tested, the size fractions applied, the shape of the respective particles, and the exposure routes tested. Our review concludes with a discussion of the important research gaps concerning freshwater ecosystems and recommendations for future areas of research.

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