Mycobacterium florentinum is a newly identified, rare, slow-growing species of nontuberculous mycobacteria (NTM). Here, we report a case of M. florentinum-induced synovitis of the wrist in an immunocompromised Japanese patient. M. florentinum was identified by sequence analysis of the rpoB, hsp65, and 16S rRNA genes. The M. florentinum strain in this study could not be differentiated from certain M. triplex strains by the hsp65 or 16S rRNA sequences alone, because they occasionally shared more than 99 % sequence identity. The isolated M. florentinum strain was only susceptible to clarithromycin and amikacin. Initially, the patient was treated with clarithromycin, levofloxacin, and ethambutol, and then with clarithromycin, levofloxacin, and rifampicin. To our knowledge, M. florentinum-induced synovitis has not been previously reported. Our results suggest that, in addition to other well-known pathogenic NTM, the recently identified M. florentinum strain should be considered as a possible cause of synovitis. Moreover, we should be cautious when identifying M. florentinum because this strain closely resembles M. triplex in genotype.