American eel (Anguilla rostrata), a once common species, is now in decline throughout much of its native range in North America. There is little information on the role of American eel in river food webs. A better understanding of the diet and ecological role of American eel will help in the conservation of this important species. During autumn 2009, eel and aquatic invertebrate samples were collected from Hannacroix Creek, a tributary of the Hudson River, in Albany and Greene counties, New York, USA. Eel diet was analyzed by the eel size and time period (day or night). A high proportion of eel stomachs were empty (73%). Eel diets varied among size classes and day and night feeding periods (p = 0.001). Diet overlap was significant between small and medium eels caught both during the day (α = 0.71) and at night (α = 0.84). Nocturnal diet and nocturnal invertebrate samples were similar (α = 0.65), indicating a preference for bottom feeding during the night. Mayfly nymphs were the major prey consumed in each period by all size classes. Among eels that fed, night-feeding eels had the greatest stomach weight (as a percent of total body weight). The swim-bladder parasite, Anguillicoloides crassus, was also observed in eels of all size classes with nearly 50% afflicted.