Identification of bifidobacterial species is still problematic because of phenotypic and genetic heterogeneities. We described a method of the Bifidobacterium identification to species based on the numerical analysis of morphological and biochemical features. Seventy five Bifidobacterium strains isolated from infants, adults, laboratory rats and bioyogurts were tested for the presence of species-characteristic cellular morphology and their biochemical patterns using the phase contrast microscopy and the API 50 CHL test, respectively. The obtained results were analysed numerically with IdBact v. 1.1 computer program (copyright by G. Kronvall, Sweden) in order to classify strains to the species. The matrices for the identification, including fermentation patterns of 20 carbohydrates, species-characteristic cellular morphology and the natural occurrence of 24 Bifidobacterium species, were created on the basis of the Bergey’s Manual identification key and the data published after 1986. PCR technique applying 16S rRNA-gene-targeted species-specific primers described by Matsuki et al. [1999] and Kok et al. [1996] were used for confirmation of the results of phenotypic identification. Type strains from American Type Culture Collection (ATCC) and Deutsche Sammlung von Mikroorganismen und Zellkulturen (DSMZ) were used as reference. The results obtained showed that bifidobacteria species significantly differ in morphology and biochemical activity. The majority of the species revealed characteristic and stable morphological traits, which in combination with their fermentation patterns enabled distinction and identification of most of the Bifidobacterium strains tested. The results of the phenotypic classification were in correlation with the performed genetical identification based on 16S rRNA-gene-targeted species-specific sequences. Finally, the tested strains were assigned to species: B. breve, B. longum, B. bifidum, B. angulatum, B. adolescentis, B. catenulatum, B. pseudocatenulatum, B. lactis, B. animalis, and B. pseudolongum.