Rapid identification of invasive Brook Trout through CRISPR‐based environmental DNA detection

Abstract Objective Invasions of nonnative fish species, such as Brook Trout Salvelinus fontinalis , disrupt ecosystem function and adversely impact native populations through competition, hybridization, and predation. Timely detection of invasive species is crucial for effective management and prevention of further spread. Environmental DNA (eDNA) technology is a powerful tool for monitoring the spread of invasive species, especially in aquatic environments, where direct observation is challenging. While eDNA sampling is sensitive and cost‐effective, real‐time analysis remains a challenge. Methods To address this limitation, we developed eDNA detection tools, using CRISPR‐Cas12a technology, that facilitate the rapid identification of invasive Brook Trout and native cutthroat trout (Lahontan Cutthroat Trout Oncorhynchus henshawi , Westslope Cutthroat Trout O. lewisi , Rocky Mountain Cutthroat Trout O. virginalis , or Coastal Cutthroat Trout O. clarkii ), without the need for specialized laboratory equipment. We employed this technology in Marsh Creek, a semi‐isolated tributary to the Sultan River in Washington State with no previous documentation of invasive Brook Trout. Result We successfully identified Brook Trout and cutthroat trout eDNA during on‐site analysis at Marsh Creek and found that the sensitivity of detecting these species can be increased with improvements in eDNA purification technology. Our results identified that Brook Trout were present in the lower reaches of the stream, particularly in areas of slow‐moving water. Conclusion The portability and simplicity of this method offer the potential for streamside eDNA detection of invasive species, and it could become another tool in the fight against the spread of invasive Brook Trout in the western United States.