Bifidobacterium adolescentis ATCC 15703, B. longum ATCC 15707, and B. thermophilum ATCC 25525 were examined for the ability to grow with fructo-oligosaccharides (FOS) as carbohydrate sources. The three species produced cell-associated β-fructosidases (inulinases) capable of hydrolysing FOS. Maximum activity was obtained with short-chain FOS with degrees of polymerization (DP) of between three and five (neosugars). The B. thermophilum inulinase was induced by inulin, a long-chain FOS with DP=35, while the enzymes from the other two strains were constitutive. Production of inulinase by all three strains was regulated by catabolite repression. Inulinase activity of the three Bifidobacterium spp. was similar when grown with 0.5% inulin as the carbohydrate source; however, B. thermophilum grew much more rapidly. All three strains utilized crude Jerusalem artichoke flour (JAF) as a carbohydrate source, suggesting that JAF might have commercial application as a food or feed additive to stimulate bifidobacteria in the gut.