Varied Pathways of Infant Gut-Associated Bifidobacterium to Assimilate Human Milk Oligosaccharides: Prevalence of the Gene Set and Its Correlation with Bifidobacteria-Rich Microbiota Formation

Abstract

The infant's gut microbiome is generally rich in the Bifidobacterium genus. The mother's milk contains natural prebiotics, called human milk oligosaccharides (HMOs), as the third most abundant solid component after lactose and lipids, and of the different gut microbes, infant gut-associated bifidobacteria are the most efficient in assimilating HMOs. Indeed, the fecal concentration of HMOs was found to be negatively correlated with the fecal abundance of Bifidobacterium in infants. Given these results, two HMO molecules, 2'-fucosyllactose and lacto-N-neotetraose, have recently been industrialized to fortify formula milk. As of now, however, our knowledge about the HMO consumption pathways in infant gut-associated bifidobacteria is still incomplete. The recent studies indicate that HMO assimilation abilities significantly vary among different Bifidobacterium species and strains. Therefore, to truly maximize the effects of prebiotic and probiotic supplementation in commercialized formula, we need to understand HMO consumption behaviors of bifidobacteria in more detail. In this review, we summarized how different Bifidobacterium species/strains are equipped with varied gene sets required for HMO assimilation. We then examined the correlation between the abundance of the HMO-related genes and bifidobacteria-rich microbiota formation in the infant gut through data mining analysis of a deposited fecal microbiome shotgun sequencing dataset. Finally, we shortly described future perspectives on HMO-related studies.

Keywords: Bifidobacterium; breast-feeding; human milk oligosaccharides; infant; microbiota.

Figure 1

HMO utilization pathways in the four major infant-gut associated Bifidobacterium species. (A–D) Degradation pathways for the representative 12 HMO molecules in B. infantis (A), B. breve (B), B. longum (C), and B. bifidum (D) are shown. The pathways in B. kashiwanohense and B. pseudocatenulatum are shown in Supplementary Figure S4. The arrows with different colors indicate the prevalence of respective homolog genes in each species (see Figure 2). Red: >75%; yellow, 25−75%; and gray: <25%. The uncharacterized degradation pathways are indicated by dotted black arrows. The extracellular enzymes and transporter homologs are shown in green and brown letters, respectively, while intracellular enzymes are in purple letters. Mono and di-saccharides liberated outside the cell membranes can be shared among gut bacteria, especially among bifidobacteria.

Figure 2

Prevalence of HMO-utilization genes in bifidobacterial genomes. Occurrence of the homolog genes (identity ≥70%, query coverage ≥60%, e value <1 × 10⁻⁵⁰) in the sixteen Bifidobacterium (sub)species genomes was examined by tblastn analysis (BLAST+ v2.9.0). Query genes used in the analysis are shown in Table 1. The prevalence (%) of the genes in each (sub)species was determined by dividing the number of the retrieved genes with the above identity criteria by the number of the genomes examined (values in parentheses). The results are shown as a heatmap.

Figure 3

Metagenomic data mining analysis of HMO-related genes of bifidobacteria. (A–C) The abundances (%) of the genes for extracellular glycosidases (A), transporters (B), and intracellular enzymes (C) detected in the metagenomic data were compared between breast-fed (BF) and formula-fed (FF) infants (≤1 year old). Data from Yatsunenko et al. [39] (n = 27 for FF and n = 34 for BF; 117,492 ± 55,056 reads/sample) were used for the analysis. See Materials and Methods for calculation of the gene abundances. Mann–Whitney U-test was used for statistical significance evaluation.

Figure 4

Heat map showing the results of Spearman’s rank correlation coefficient analysis between the relative abundance of genus Bifidobacterium and the relative abundance of HMO (A) or GOS utilization genes (B). The fecal metagenomic data of the infants living in USA, Malawi, and Venezuela were used for the analysis. The abundance of the genus Bifidobacterium was obtained from MG-RAST version 4.0.3 (see Material and Methods). See also, Supplementary Figure S5 for HMO degradation and transport genes (A) and Supplementary Figure S6 for GOS transporter genes (B). *, **, ***, and **** denote p < 0.05, p < 0.01, p < 0.001, and p < 0.0001 respectively. BF: breast-fed; FF: formula-fed; GL-BP: galactosyllactose-binding protein; N.A.: correlation analysis was not applicable due to the absence of the reads attributable to FL1/2-BP or GlcNAc-ase (Blon_2355) genes in the tested subjects.