A numerical analysis based on phenotypic characteristics (89 enzymatic tests and 49 carbohydrate acidification tests), in which experimental strips from Biomerieux-API, La Balme les Grottes, France, were used, was performed to characterize 82 new isolates belonging or related to Bijidobacterium longum, Bifidobacteriurn infantis, and Bafuiobucteriurn breve. A total of 72 strains were isolated from child or adult feces, and the other strains were obtained from human vaginas and bronchi. In this study we also included 38 type and reference strains that were representative of all species of the genus Bifidobucterium and 6 strains belonging to the genus LactobuciUus, DNA-DNA relationships between B. longurn and B. infantis were determined by using 19 strains related to these species, as determined by the numerical analysis, The degree of DNA binding was determined by the S1 nuclease method. The phenotypic study revealed that there were six main clusters, which were subdivided into nine subclusters. Subcluster Va contained the type strains of B. longum and B. infantis. The DNA-DNA relatedness values of some of the new isolates were very similar to the DNA-DNA relatedness values of the type strain of B. longurn. On the basis of these data, it was difficult to isolate B. infantis strains and then to define B. infantis as a single species separated from B. longurn. Subclusters lVb to IVf comprised reference strains of B. breve. Cluster I11 and subcluster Ia were not identified.At the species, present time, the genus Bifidobacterium includes 29 10 of which are of human origin. Four of these species, Bijidobacterium bijidum, Bijidobacterium longum, Bifidobacterium infantis, and Bifidobacterium breve, have been studied with increasing interest because of their role in physiological relationships in human (especially infant and child) gastrointestinal tracts (2,3,29). Furthermore, today, the use of these bacteria in food microbiology (dairy products) makes accurate differentiation of these organisms necessary (19).The type species of the genus, B. bifidum, is well separated from the other species on the basis of its phenotypic characteristics, as well as its DNA-DNA hybridization values. Identification of the three other species studied in this work is much more uncertain; in particular, distinction between B. longum and B. infantis is difficult.On the basis of the results of phenotypic studies, Reuter (18) defined and subdivided B. longum into two biovars, biovars a and b, and Mitsuoka (17)

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