Tuberculosis is still the greatest single infectious cause of mortality in the world. Moreover, the spread of HIV has increased the number of tuberculosis patients, yet powerful new anti-TB drugs with new mechanisms of action have not been developed in over thirty years, with only first-line antituberculous drugs still in use. We screened for new antimycobacterial antibiotics from microbial products with effective and specific narrow-range spectra, and as part of this program, discovered a novel anti-tuberculosis antibiotic named caprazamycin B (CPZ-B) from the culture broth of the Actinomycete strain Streptomyces sp. MK730-62F2. CPZ-B is a new lipo-nucleoside antibiotic related to liposidomycins and capuramycin, with a molecular formula of C53H86N5O22. It showed excellent in vitro anti-mycobacterial activity against drug-susceptible and multidrug-resistant M. tuberculosis strains: MICs were 3.13 μg/ml for M. tuberculosis H37Rv and Kurono strains, 3.13 μg/ml for M. bovis Ravenel strain, 6.25–12.5 μg/ml for drug-susceptible M. tuberculosis (n=22), and 6.25–12.5 μg/ml for multidrug-resistant M. tuberculosis (n=12). It also exhibited in vitro activity equivalent to clarithromycin against Mycobacterium avium complex (MAC), with MICs of 6.25–50 μg/ml for M. avium (n=33) and 1.56–25 μg/ml for M. intracellulare (n=17). CPZ-B demonstrated no significant toxicity in mice (single dose >200 mg/kg i.v., repeated dose 100 mg/kg for 14 days) and no genotoxicity or cytotoxicity (5000 μg/ml). Thus, CPZ-B is considered a promising candidate for an anti-TB drug.