Prebiotics are non-digestible substrates that stimulate the growth of beneficial microbial populations in
the intestine, especially Bifidobacterium species. Among them, fructo- and galacto-oligosaccharides are
commonly used in the food industry, especially as a supplement for infant formulas. Mechanistic details
on the enrichment of bifidobacteria by these prebiotics are important to understand the effects of these
dietary interventions. In this study the consumption of galactooligosaccharides was studied for 22
isolates of Bifidobacterium longum subsp. infantis, one of the most representative species in the infant gut
microbiota. In general all isolates showed a vigorous growth on these oligosaccharides, but consumption
of larger galactooligosaccharides was variable. Bifidobacterium infantis ATCC 15697 has five genes
encoding b-galactosidases, and three of them were induced during bacterial growth on commercial
galactooligosaccharides. Recombinant b-galactosidases from B. infantis ATCC 15697 displayed different
preferences for b-galactosides such as 40 and 60-galactobiose, and four b-galactosidases in this strain
released monosaccharides from galactooligosaccharides. Finally, we determined the amounts of short
chain fatty acids produced by strain ATCC 15697 after growth on different prebiotics. We observed that
biomass and product yields of substrate were higher for lactose and galactooligosaccharides, but the
amount of acids produced per cell was larger after growth on human milk oligosaccharides. These results
provide a molecular basis for galactooligosaccharide consumption in B. infantis, and also represent
evidence for physiological differences in the metabolism of prebiotics that might have a differential
impact on the host.