The identification of predatory animals is essential to reducing fisheries damage. In this study, we investigated the presence of fish genes in the excrement of Great Cormorant Phalacrocorax carbo at a reservoir in Gifu Prefecture, Japan. DNA metabarcoding analysis revealed that most fish species eaten by Great Cormorants were Japanese Barbel Hemibarbus barbus (number of birds detected/tested, 10/16), Japanese Dace Pseudaspius hakonensis (9/16), and Japanese Smelt Hypomesus nipponensis (9/16), whereas the genes of released fish were detected in only a few samples. Our findings suggest that cormorants living at the reservoir rarely consumed released fish.