Stable isotope analysis (SIA) was used to quantify intra- and interspecific variation in the δ¹³C and δ¹⁵N values in plagiopatagium tissue of little brown (Myotis lucifugus) and northern long-eared (M. septentrionalis) bats in several regions of the Canadian Maritimes where they occur in sympatry. There was large intraspecific variation in the δ¹³C of M. lucifugus, with the range exceeding 30‰, whereas the range of δ¹³C values observed in M. septentrionalis was less than 7‰. Similarly, the standard ellipse area of M. lucifugus was larger than that of M. septentrionalis at all sites and together, these data support the contention that M. lucifugus has broader dietary niche breadth than M. septentrionalis. Some M. lucifugus from Fundy National Park, New Brunswick exhibited very low δ¹³C values, suggesting an energy input from an unknown source, possibly with carbon assimilated from biogenic methane. High δ¹³C values for M. lucifugus from Brier Island, Nova Scotia are consistent with a diet that is at least partially derived from marine sources. Finally, δ¹⁵N values for both species from Prince Edward Island were high relative to New Brunswick or Nova Scotia, suggesting inputs of anthropogenically-derived nitrogen from a more agriculturally-intensive landscape.