The Cuban treefrog (Osteopilus septentrionalis) is an introduced amphibian that has successfully established widespread populations across Puerto Rico. Despite its invasive status and potential impacts on native species, genetic information for local populations remains limited. This project aims to genetically characterize Puerto Rican populations of O. septentrionalis using mitochondrial COI DNA barcoding obtained exclusively through non-invasive sampling methods. The resulting data will support taxonomic validation and improve understanding of genetic variability, contributing to future research and management of invasive amphibian species.