Metabolic cross-feeding relationship gains probiotic status

However, even in the gut environment where these molecules are highly abundant, species have been identified that are unable to digest the sometimes highly complex plant polymers. This means these species depend on other members of the community to break down their food for them – called ‘syntrophy’. In the study by Muñoz et al. discussed here, a novel syntrophic relationship was identified in which Bacteroides cellulosilyticus breaks down side chains of a highly complex plant peptidoglycan. Bifidobacterium breve can then use these degradation products as sole carbon sources for growth. As such, the list of probiotic organisms is being extended by a syntrophic relationship while more plant components gain the status of prebiotics.

Unprocessed plant products contain the complex and sturdy molecules that make up the plant cell wall, which our digestive enzymes are unable to deal with. Generally, plant polymers like cellulose, hemicellulose or lignin are meant to stabilise and protect the plant from physical breakage and enzymatic digestion. One such polymer that is omnipresent in plant cell walls is an extracellular proteoglycan called arabinogalactan protein (AGP). Almost all vegetal foods contain AGPs, with gum arabic (Arabic gum) from the Acacia senegal tree being the best known and most widely used natural gum. The food industry discovered the emulsifying properties of AGPs and use them in fruit syrups, marshmallows, confectionary sugar, icings, chewing gum, soft drinks and in edible decorative ingredients. Due to the vital roles AGPs play in our diet and the incompetence of the human digestive enzymes to degrade this polymer, it is of utmost importance to understand the fate of AGPs in our guts.

While every plant seems to produce its own unique AGP, they are thought to be involved in many physiological events. Interestingly, only a small fraction of AGPs are actually peptide content – even though being rich in hydrophilic amino acids is what gives AGPs their characteristic high solubility. Over 90% of AGPs are made up of carbohydrates, commonly referred to as arabinogalactans (AGs), which are neutral or slightly acidic complex polysaccharides with a varying number of side chains. The main chain consists of 1,3-linked β-d-galactopyranosyl units, which branch via 1,6-linkages into side chains. These side chains have variable additional sugars, linkages and lengths and are often capped by α-l-rhamnose sugars.

When glycan components are not digested by human enzymes, they become available to the human gut microbiota, which after anaerobic fermentation uses the building blocks as nutrients. Many members of the human gut microbiota encode carbohydrate-active enzymes (CAZymes) able to ferment complex glycans. For example, Bacteroides thetaiotaomicron, a commensal member of the human gut microbiota, encodes 288 glycoside hydrolases, which cleave the glycosidic bonds linking the sugars in polysaccharides and oligosaccharides. Furthermore, in many Bacteroides genomes, so-called polysaccharide utilisation loci (PULs) have been identified, consisting of CAZymes, glycan transporters and sensors for transcription regulation.

Several Bacteroides species (such as B. thetaiotaomicron, B. cellulosilyticus and Bacteroides finegoldii) have been shown to produce extracellular enzymes that cleave off the terminal rhamnose molecule and the 1,6-linked side chains from AGs. The Bacteroides bacteria would start depolymerising the backbones of the AGs and release the broken oligosaccharides into the medium. Other members of the commensal microbiota are then thought to pick up and subsequently degrade the released oligosaccharides. It has been generally assumed that these bacteria lack the enzymes to degrade the highly complex AGs and require the first degradation step by the Bacteroides for their survival.

A novel study by Muñoz et al. showed that the commensal Bifidobacterium breve could metabolise oligosaccharides released by B. cellulosilyticus from AG degradation. B. breve was previously shown to metabolise human milk oligosaccharides and dietary glycans, but seems to be unable to digest complex oligosaccharides like the highly complex AG found in Arabic gum. The authors showed that B. breve contains a 3 gene cluster encoding for a transcriptional regulator, a sugar symporter and the glycoside hydrolase-like enzyme BgaA. BgaA was shown to exhibit hydrolytic activity towards different 1,3-oligosaccharides. A deletion mutant in this enzyme showed notable growth defects when grown on these oligosaccharides and it lost its ability to grow in co-culture with B. cellulosilyticus. Hence, BgaA can be understood as a vital factor for B. breve growth as it renders B. breve able to utilise oligosaccharides produced by B. cellulosilyticus.

Lastly, the authors aimed to show the prebiotic nature of AGs by analysing the release of short chain fatty acids (SCFAs) after the syntrophic metabolic activity of both strains. It was previously revealed that B. cellulosilyticus produces succinate and acetate when grown on AGs. The authors showed that when B. breve and B. cellulosilyticus grew in co-culture on AGs, the SCFA levels increased due to the syntrophic behaviour. They also found that thanks to the cross-feeding process, B. breve produces acetate, lactate and formate, thus rendering SCFA production higher and more diverse. Due to the production of SCFAs, this syntrophic interaction between B. breve and B. cellulosilyticus has probiotic character, while AGs have now gained the status of prebiotics.

Plant cells contain other rigid polymers that are subject to syntrophic interactions within the human gut microbiota, for example xylan. This polymer consists of β-(1,4)-linked xylose residues substituted with various sugar derivates or acetyl groups. When grown on complex xylan, Bacteroides ovatus was shown to support the growth of Bifidobacterium adolescentis, which can only utilise linear arabino-xylo-oligosaccharides released by B. ovatus. Similarly, B. breve and Escherichia coli metabolise fucose and rhamnose, released from plant polymers, into 1,2-propanediol. This component has an enhancing growth effect on the probiotic Lactobacillus reuteri. Subsequently, from 1,2-propanediol, L. reuteri and Eubacterium hallii produce propanol and propionate, also key SCFAs with important health benefits.

Identifying another syntrophic relationship that is active in the human gut microbiota drew more and more attention to the commonly used phrase ‘You are what you eat’. As discussed here, Muñoz et al. identified the probiotic character of a new syntrophic relationship and the prebiotic nature of AGs. However, we are still far from completely understanding the metabolic relationships and cross-feeding networks going on within the gut microbiota. But we can be assured that there are many more advantageous metabolic avenues of probiotics yet to be discovered.