Freshwater ecosystems are negatively impacted by various pollutants, from agricultural, urban and industrial wastewater, with metals being one of the largest concerns. Moreover, freshwater ecosystems are often affected by alien species introductions that can modify habitats and trophic relationships. Accordingly, the threat posed by metals interacts with those by alien species, since the latter can accumulate and transfer these substances across the food web to higher trophic levels. How metals transfer within such communities is little studied. We analysed the concentration of 14 metals/metalloids (Al, As, Cd, Co, Cr, Cu, Fe, Hg, Mg, Mn, Ni, Pb, Se, Zn, hereafter 'metal(s)') of eight fish and three crustacean species co-existing in the Arno River (Central Italy), most of which were alien. To assess the pathway of contaminants within the community, we coupled metal analysis with carbon and nitrogen stable isotope analysis derived from the same specimens. Crustaceans showed higher metal concentration than fish, except for Cd, Hg and Se that were higher in fish. We found evidence of trophic transfer for six metals (Cd, Cr, Hg, Mg, Se, Zn). Additionally, ontogenetic differences and differences among various fish tissues (muscle, liver, and gills) were found in metals concentration. Considerable biomagnification along the trophic chain was found for Hg, while other metals were found to biodilute. Using stable isotopes and Hg as a third diet tracer, we refined the estimations of consumed preys in the diet previously reconstructed with stable isotope mixing models. Alien species reach high biomass and can both survive to and accumulate high pollutants concentrations, potentially posing a risk for their predators and humans. A combined effect of environmental filtering and increased competition may potentially contribute to the disappearance of native species with lower tolerances.

Senckenberg Research Institute and Natural History Museum Frankfurt Department of River Ecology and Conservation Clamecystrasse 12 63571 Gelnhausen Germany

University of Duisburg Essen Faculty of Biology Universitätsstraße 5 45141 Essen Germany

University of Florence Department of Biology Via Madonna del Piano 6 50019 Sesto Fiorentino Italy

University of South Bohemia in České Budějovice Faculty of Fisheries and Protection of Waters South Bohemian Research Center of Aquaculture and Biodiversity of Hydrocenoses Zátiší 728 2 389 25 Vodňany Czech Republic

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