| Summary: | The development of infant gut microbiota is strongly influenced by nutrition. Human milk oligosaccharides (HMOSs) in breast milk selectively promote the growth of glycan-degrading microbes, which lays the basis of the microbial network. In this study, we investigated the trophic interaction between <i>Bacteroides thetaiotaomicron</i> and the butyrate-producing <i>Anaerostipes caccae</i> in the presence of early-life carbohydrates. Anaerobic bioreactors were set up to study the monocultures of <i>B. thetaiotaomicron</i> and the co-cultures of <i>B. thetaiotaomicron</i> with <i>A. caccae</i> in minimal media supplemented with lactose or a total human milk carbohydrate fraction. Bacterial growth (qPCR), metabolites (HPLC), and HMOS utilization (LC-ESI-MS<sup>2</sup>) were monitored. <i>B. thetaiotaomicron</i> displayed potent glycan catabolic capability with differential preference in degrading specific low molecular weight HMOSs, including the neutral trioses (2′-FL and 3-FL), neutral tetraoses (DFL, LNT, LNnT), neutral pentaoses (LNFP I, II, III, V), and acidic trioses (3′-SL and 6′-SL). In contrast, <i>A. caccae</i> was not able to utilize lactose and HMOSs. However, the signature metabolite of <i>A. caccae</i>, butyrate, was detected in co-culture with <i>B. thetaiotaomicron</i>. As such, <i>A. caccae</i> cross-fed on <i>B. thetaiotaomicron</i>-derived monosaccharides, acetate, and <span style="font-variant: small-caps;">d</span>-lactate for growth and concomitant butyrate production. This study provides a proof of concept that <i>B. thetaiotaomicron</i> could drive the butyrogenic metabolic network in the infant gut.
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