Evaluating eDNA Detection from Live and Dead Control Sources

Environmental DNA (eDNA) methods offer a powerful tool for monitoring aquatic species, yet field applications remain challenged by uncertainty in DNA transport, mixing, and detection, particularly in flowing or tidally influenced systems. One approach to improve confidence in eDNA surveys is the use of controlled DNA sources (positive controls), but questions remain regarding how the biological condition of the source influences eDNA release and detectability. This study evaluated differences in eDNA concentrations emitted from live versus dead fish in a controlled, shallow, well-mixed channel. Using a fixed point-sampling design, we measured eDNA concentrations over time and modeled the effects of treatment, sampling time, temperature, and water velocity. Dead fish consistently released significantly higher concentrations of eDNA than live fish, while eDNA concentrations declined over time in both treatments. Water temperature and velocity did not significantly influence detection, and the rate of eDNA decline was similar between live and dead treatments. These findings highlight the importance of source condition and site-specific mixing dynamics when interpreting positive control experiments and underscore the value of site characterization when designing eDNA sampling protocols.