Vitamin B 12 is a complex biomolecule that acts as a cofactor for a variety of enzymes in microbial metabolism. Pseudomonas denitrificans is exclusively used as an industrial strain for the production of vitamin B 12 under aerobic conditions. However, only a few strains of Pseudomonas have been reported to possess the capability of producing this vitamin and they are strongly patentprotected. To improve the applicability of the vitamin B 12producing microorganisms, a new isolate was obtained from municipal waste samples and characterized for its biological properties. The new isolate, designated as SP2, was identified to be a Pseudomonas species based on the sequence homology of its 16S rDNA. Pseudomonas species SP2 had essential genes for vitamin B 12 synthesis such as cobB and cobQ and produced a similar amount of vitamin B 12 (10.6 ± 0.05 µg/mL) as P. denitrificans ATCC 13867 in 24 h flask culture. SP2 grew well under aerobic condition with the maximum specific growth rate (µ max ) of 0.91 + 0.03/h, but showed a poor growth under micro-aerobic conditions. SP2 was resistant to antibiotics like streptomycin, carbenicillin, ampicillin, cefpodoxime, colistin, nalidixic acid and sparfloxacin. The ability of SP2 to grow faster and produce vitamin B 12 under aerobic conditions makes it a promising host for the production of some biochemicals requiring a coenzyme B 12 -dependent enzyme, such as glycerol dehydratase.