A total of twenty-two strict anaerobic and Gram-positive Bifidobacteria, identified as B. adolescentis, B.pseudocatenulatum, or B. longum, were isolated from healthy adult Koreans. We here investigated the cell morphology, antimicrobial resistance patterns to novel antibiotics and genotypic differentiation of Bifidobacteria assessing repetitive DNA element PCR (rep-PCR) fingerprinting using the BOXA1R primer at the species level. All Bifidobacterium spp., except B.adolescentis SPM1005 and B. longum SPM1205, formed round and convex colonies. All B. adolescentis, B. pseudocatenulatum, and B. longum were opaque white glossy in colony color, and short, long, and irregular rods in morphological shape. In addition, all B.adolescentis, B. pseudocatenulatum, and B. longum formed a variety of shapes ranging from rods to Vshaped, Y-shaped, clubbed rods, or irregular. All Bifidobacterium spp., except B. adolescentis SPM0214, were sensitive to daptomycin (DAP), linezolid (LIN), and tigecycline (TIG). B. adolescentis SPM0214 was resistant to DAP. Genomic fingerprinting patterns of B. adolescentis, B. pseudocatenulatum, and B. longum were diverse and different from those of the KCTC strain. The band size of B. adolescentis, B.pseududocatenulatum, and B. longum varied from 3.0 kb to 300 bp, 2.0 kb to 200 bp, and 2.0 kb to 500 bp, respectively. In conclusion, twenty-two strains of B.adolescentis, B. pseudocatenulatum, and B. longum isolated from healthy adult Koreans were very diverse in both phenotype and genotype. Moreover, this diversity of phenotype and genotype may support that health promoting effects of individual strain of Bifidobacterium spp. human isolates could be different and specific even within same species.