Effective recovery plans for endangered species rely on insights into species’ ecology to identify risks and develop population recovery strategies. Data gaps pose challenges for many species of conservation concern, particularly those with cryptic behaviors or that occupy difficult-to-access habitats. Sounds produced by these species offer an effective means of observing many such marine and aquatic species, and for this reason, passive acoustic monitoring has emerged as an important study and assessment approach in marine systems. This approach is only just beginning to be applied for aquatic species monitoring in freshwater habitats. Atlantic sturgeon Acipenser oxyrinchus, a species of conservation concern along the US East Coast, remains poorly understood due to persistent data gaps despite years of conservation efforts. While sounds have been described for other sturgeons, sounds from Atlantic sturgeon have not yet been reported. Here, we characterized acoustic cues associated with Atlantic sturgeon in the Hudson River, New York, USA, and identified a low-frequency (44 Hz peak frequency) signal strongly correlated with the occurrence of telemetry-tagged adults which enter the river to spawn. We corroborated these efforts with recordings of captive Atlantic sturgeon, in which we detected the same sound type during a spawning period. Our findings provide an opportunity to develop passive acoustic monitoring strategies for Atlantic sturgeon, offering a non-invasive tool for understanding the spatiotemporal distribution of spawning activity across their range. We demonstrate potential applications of passive acoustic monitoring to inform sturgeon conservation and management, including characterizing habitat use, identifying cross-species interactions, and providing abundance indices.