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The spectrum size has been used to assess environmental disturbances and to understanding the energy flow in ecosystems. The objective of this work was to investigate if the operation regimes of reservoirs, including run-of-river and storage systems, interfere with the biomass spectra of fish fauna. We tested the hypothesis that the run of river reservoirs present higher proportions of large individuals than storage system. Samplings of fish fauna were carried out between January 2005 and December 2007 at six Neotropical reservoirs belonging to the sub-basin Iguaçu River and Coastal basin in São Jorge River, Brazil. The spectrum calculation was performed using the Pareto type I continuous distribution model. Reservoirs operated under run-of-river regime had significantly higher values than those operated under storage regimes. This study has elucidated some impacts of reservoir operating regime on biomass spectra and indicated differences in size spectra of fish assemblages among the sampled reservoirs. Therefore, it is important to incorporate management plans that take dam operating mode into account so that conservation of aquatic fauna, especially fish, is more effective.