This study examined mudpuppy populations in the St. Clair-Detroit River System using trapping data, environmental DNA (eDNA) sampling, and land cover analysis. The objectives were to determine if eDNA is a useful monitoring tool for mudpuppies and quantify the relationship between land cover and mudpuppy presence. The results showed that sites with higher percentages of forest cover and lower development upstream had higher mudpuppy occupancy. eDNA results were inconclusive for monitoring. Ongoing sampling and protocol adjustments may help validate the use of eDNA to monitor mudpuppy populations.