Mycobacterium tuberculosis is responsible for Tuberculosis (TB), which is the deadliest infectious disease in human history. Concerted investment in TB research has guided healthcare interventions that have resulted in a 2% annual reduction in global TB incidence and mortality since 1990. In contrast, opportunistic non-tuberculous mycobacteria (NTM) infections are increasing worldwide, even among populations served by the world’s most advanced healthcare systems. The simultaneous opposing trends of TB and NTM disease incidence implies a knowledge gap between the evolutionary ecology of transmissible and environmental mycobacterioses . We sought to narrow this gap by investigating an aquatic NTM species recently reported from the Neotropics , including the northern Gulf of Mexico. Mycobacterium ulcerans is responsible for Buruli Ulcer (BU), a necrotic infection of skin and soft tissues that is of substantial public health concern in tropical regions of the eastern hemisphere. The same bacterium is also responsible for disease outbreaks in global aquaculture, resulting in economic losses exceeding $1m annually. We surveyed cultured and wild populations of sportfishes , baitfishes , and invertebrates from Louisiana, Mississippi, Alabama, Florida, and Puerto Rico. Using molecular detection methods, we found evidence for diverse and widespread lineages of M. ulcerans , including strains known to cause debilitating human disease such as BU. Molecular sequence data was used to infer a new phylogeny of the M. ulcerans complex, which reveals evidence for intercontinental translocation of lineages of importance in aquaculture. Though mycobacterial disease in fishes of the Gulf of Mexico appears to be rare, we caution that the presence of this zoonotic pathogen imparts a need for integrative risk assessment among stakeholders from aquaculture and public health organizations.