Novel Genes and Metabolite Trends in Bifidobacterium longum subsp. infantis Bi-26 Metabolism of Human Milk Oligosaccharide 2′-fucosyllactose

Abstract

Human milk oligosaccharides (HMOs) function as prebiotics for beneficial bacteria in the developing gut, often dominated by Bifidobacterium spp. To understand the relationship between bifidobacteria utilizing HMOs and how the metabolites that are produced could affect the host, we analyzed the metabolism of HMO 2′-fucosyllactose (2′-FL) in Bifidobacterium longum subsp. infantis Bi-26. RNA-seq and metabolite analysis (NMR/GCMS) was performed on samples at early (A600 = 0.25), mid-log (0.5–0.7) and late-log phases (1.0–2.0) of growth. Transcriptomic analysis revealed many gene clusters including three novel ABC-type sugar transport clusters to be upregulated in Bi-26 involved in processing of 2′-FL along with metabolism of its monomers glucose, fucose and galactose. Metabolite data confirmed the production of formate, acetate, 1,2-propanediol, lactate and cleaving of fucose from 2′-FL. The formation of acetate, formate, and lactate showed how the cell uses metabolites during fermentation to produce higher levels of ATP (mid-log compared to other stages) or generate cofactors to balance redox. We concluded that 2′-FL metabolism is a complex process involving multiple gene clusters, that produce a more diverse metabolite profile compared to lactose. These results provide valuable insight on the mode-of-action of 2′-FL utilization by Bifidobacterium longum subsp. infantis Bi-26.

Strain-specific strategies of 2′-fucosyllactose, 3-fucosyllactose, and difucosyllactose assimilation by Bifidobacterium longum subsp. infantis Bi-26 and ATCC 15697

“…Although beyond the scope of this study, this phenomenon warrants further investigation, as fucose metabolism is highly relevant in vivo in the mammalian host 61,62 , an environment where fucose is an abundant component of mucin and milk glycans and glycoproteins, potentially available to gut microbiota 63,64 and is one of important mediators of host-microbe symbiosis 61,[64][65][66][67][68] . Pyruvate excretion by B. infants during growth on HMOs has not been reported in the literature, except as a brief mention in an earlier publication from our group 24 . In addition to pyruvate excretion by Bi-26 ( Fig.…”

“…The fate of the remainder of the fucosyl moiety in the type strain or in Bi-26 was not pursued in this study. However, from out earlier work 24 we know that at least a part of it is excreted by Bi-26 as L-fucose during growth on 2′FL 24 and that the amounts of 1,2-PD and L-fucose accumulating in the spent medium inversely correlated under these conditions (not shown). In the absence of any fucose catabolic pathways in Bifidobacterium genera other than the non-phosphorylated anaerobic route, the L-fucose monosaccharide excretion and its catabolism to pyruvate and 1,2-PD are the only two alternative fates of the fucosyl moiety in FLs (or other fucosylated HMOs) conceivable to date (illustrated schematically in Fig.…”

“…longum, and B. kashiwanohense the capacity for uptake and catabolism of HMOs is limited to specific strains and to selected types of HMOs 18,30,51 , in B. infantis strong growth on a wide range of HMOs is considered characteristic of the entire subspecies 20,52,53 . The detailed analysis of molecular mechanisms of HMO utilization has been addressed largely in a single representative of B. infantis subspecies, the type strain ATCC 15697, with a few exceptions 20,24 leading to the underestimation of the genetic and metabolic diversity present within the B. infantis subspecies as a whole. In this study we show that at least two distinct strategies for FL utilization are evident within the B. infantis subspecies.…”

“…Genes involved in recognition and initial processing of 2′FL, 3FL, or DFL. Since we are primarily interested in HMO utilization, we focused on genes known to be involved in HMO utilization in the type strain [19][20][21][22]29 and other bifidobacteria 24,25,30 to look for differences between the two strains tested. The gene list is shown in Figs.…”

Influence of 2′-fucosyllactose and galacto-oligosaccharides on the growth and adhesion of Streptococcus mutans

“…B. longum subsp. infantis has several glycoside hydrolases, ATP binding cassette (ABC) transporters and extracellular solute binding proteins, it utilises to import HMOs into the cell and degrade oligosaccharides intracellularly (39)(40)(41) . Bacteroides, on the other hand, have been proposed to degrade complex HMOs partly and then transport processed oligosaccharides inside the cell for further degradation (39) .…”