The Indian River Lagoon is one of the most species-rich estuaries in the United States, providing habitat to over 400 species of fish. Among these are critical forage fishes, such as menhaden, anchovy, and sardines, as well as many of their commercially important predators. The Florida Fish and Wildlife Conservation Commission (FWC) conducts monthly seine surveys in the IRL, which provide the basis for most fisheries resource management and conservation decisions in the region. However, many key species are systematically overlooked by these surveys due to gear bias, resulting in data deficiencies for many forage fish species and their dependent predators. This ongoing study aims to circumvent these issues by utilizing environmental DNA (eDNA) metabarcoding to conduct a survey of forage fish species in the IRL and create a rapid and cost-effective survey toolkit complimenting existing survey efforts. As part of this effort, we have collected water samples from 16 sites across the northern IRL alongside FWC’s monthly surveys just before seine net deployment. Using protocols optimized in our lab at the University of Central Florida, we will extract DNA from these samples and prepare Illumina libraries using 16S primers. Species composition and individual occurrence records at each sample site and habitat will be evaluated based on the eDNA data. These results will be compared to those obtained from the FWC survey to evaluate the relative strengths and weaknesses of each method. Combining the data, we will create local and regional biodiversity hotspot maps for forage fishes that can be used to supplement FWC’s species occurrence database.