Abstract Recent surveys of periphyton in Lake Tikitapu revealed widespread benthic mats dominated by cyanobacteria. All mats tested positive for the cyanobacterial toxin nodularin-R. The New Zealand native freshwater crayfish or kōura (Paranephrops planifrons) are benthic-dwelling, opportunistic omnivores that are common in Lake Tikitapu. Benthic mats constitute a potential food source for this species. In this study an in-lake feeding experiment with isotopically labeled 13C benthic mats confirmed they were consumed by kōura. Consumption was variable amongst individuals, suggesting the benthic mats are an optional rather than primary food source. Nine kōura were also tested using liquid chromatography–mass spectrometry to determine if nodularin-R bioaccumulated in the hepatopancreas and tail tissue. The hepatopancreas of all kōura were positive for nodularin-R (9.7–225.3 μg kg−1 ww) and nodularin-R was detected in low concentrations in the tail tissue of two individuals (0.5–0.7 μg kg−1 ww). The detection of nodularin-R in kōura is the first in a freshwater organism in a freshwater system, and the first to show the accumulation of nodularin-R from freshwater benthic cyanobacterial mats. Benthic mats may need to be considered as a potential source of cyanotoxins in future freshwater food-web studies.