Seagrass conservation is critical for mitigating climate change due to the large stocks of carbon they sequester in the seafloor. However, effective conservation and its potential to provide nature-based solutions to climate change is hindered by major uncertainties regarding seagrass extent and distribution. Here, we describe the characterization of the world’s largest seagrass ecosystem, located in The Bahamas. We integrate existing spatial estimates with an updated empirical remote sensing product and perform extensive ground-truthing of the seafloor with 2,542 diver surveys across remote sensing tiles. We also leverage seafloor assessments and movement data obtained from instrument-equipped tiger sharks, which have strong fidelity to seagrass ecosystems, to augment and further validate predictions. We report a consensus area of at least 66,000 km2 and up to 92,000 km2 of seagrass habitat across The Bahamas Banks. Sediment core analysis of stored organic carbon further confirmed the global relevance of the blue carbon stock in this ecosystem. Data from tiger sharks proved important in supporting mapping and ground-truthing remote sensing estimates. This work provides evidence of major knowledge gaps in the ocean ecosystem, the benefits in partnering with marine animals to address these gaps, and underscores support for rapid protection of oceanic carbon sinks.