Fatty acids (FA) have recently been used in several studies to infer the diet in a number of species. While these studies have been largely successful, most have dealt with predators that have a fairly specialized diet. In this paper, we used FA analysis as a tool to infer the diet of the nearctic river otter (Lontra canadensis). The river otter is an opportunistic predator known to subsist on a wide variety of prey including, fishes, crayfish, molluscs, reptiles and amphibians, among others. We analyzed the principle components of 60 FA from otters and 25 potential prey species in Illinois, USA. Prey species came from 4 major taxonomic divisions: fishes, crayfish, molluscs and amphibians. Within each division, most, but not all, species had significantly different profiles. Using quantitative FA signature analysis, our results suggest that, by mass, fish species are the most significant component of Illinois River otters’ diet (37.7 ± 1.0%). Molluscs ranked second (32.0 ± 0.8%), followed by amphibians (27.3 ± 4.3%), and finally, crayfish (3.0 ± 0.6%). Our analysis indicates that molluscs make up a larger portion of the otter diet than previously reported. Throughout much of the Midwest there have been numerous otter reintroduction efforts, many of which appear to be successful. In regions where mollusc species are endangered, these data are essential for management agencies to better understand the potential impact of otters on these species. Our analysis further suggests that quantitative FA signature analysis can be used to infer diet even when prey species are diverse, to the extent that their FA profiles differ. Better understanding of the otter’s metabolism of FA would improve inferences of diet from FA analysis.
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