Abstract: Mycobacteriosis in fish is a common chronic progressive disease spread worldwide. Several outbreaks of mycobacteriosis in reared sturgeons occurred in China in year 2009 and 2010, which affected Chinese sturgeon (Acipenser sinensis), Amur sturgeon (Acipenser schrencki) and hybrid sturgeon (Acipenser baeri-Acipenser gueldenstaedtii). Mycobacteria were identified as the pathongens of this disease based on a variety of observation, including the identification of the isolates, the detection of pathogens in tissues, the clinical signs and the histopathological examinations. Forty-nine isolates of nontuberculous mycobacteria were extracted from 19 infected sturgeons. We identified seven species of Mycobacterium from these isolates, namely, Mycobacterium chelonae, Mycobacterium marinum, Mycobacterium gordonae, Mycobacterium fortuitum, Mycobacterium szulgai, Mycobacterium porcinum, and Mycobacterium arupense. We found that two or three mycobacterial species co-existed in the same tissues. Our results also showed that M. marinum was more prevalent than other mycobacterial species. Virulence assay revealed that M. marinum showed the highest pathogenicity to zebrafish. These results suggested that M. marinum should be the major pathogenic bacteria in sturgeon mycobacteriosis, and that the mixed mycobacterial infection be the predominant form of sturgeon mycobacteriosis. To our best knowledge, this is the first report about the mycobacteriosis in farmed Chinese sturgeons and Amur sturgeons, and is the first report about the isolation of M. porcinum and M. arupense from the infected fish.