Mediterranean freshwaters undergo extreme seasonal variation in water flow, which, exacerbated by water withdrawal for agriculture or hydroelectric purposes, may affect fish communities and thus prey availability for semi-aquatic predators, such as Eurasian otter Lutra lutra. To investigate the role played by food availability on the ongoing recovery of an otter population at the southernmost limit of its Italian range, we assessed otter diet by the analysis of 357 spraints collected from 2014 to 2017 on eight rivers, and both fish and amphibian availability by, respectively, electrofishing and visual encounter surveys. Fish and amphibians formed the bulk of otter diet, the latter resource contributing as much as fish to otter diet in spring. Use by otters of both fish and amphibians depended only fish availability, suggesting that amphibians constituted an alternative resource to be exploited in conditions of fish shortage. Accordingly, electrofishing showed that fish biomass may barely be sufficient to sustain the current otter population. ATR-FT-IR spectroscopy allowed to point out for the first time the occurrence of amphibian eggs in otter spraints, although the co-occurrence of anuran bones did not allow to discriminate between direct and passive predation. Overall results indicate that the expansion or even survival of this small otter population may depend on the effective management of water resources and reinforcement of fish assemblages.