Sharing a β-Glucan Meal: Transcriptomic Eavesdropping on a Bacteroides ovatus-Subdoligranulum variabile-Hungatella hathewayi Consortium

Abstract

Whole-transcriptome analysis was used to investigate the molecular interplay between three bacterial species that are members of the human gut microbiota. Bacteroides ovatus, Subdoligranulum variabile, and Hungatella hathewayi formed associations in cocultures fed barley β-glucan, a constituent of dietary fiber. B. ovatus depolymerized β-glucan and released, but did not utilize, 3-O-β-cellobiosyl-d-glucose (DP3) and 3-O-β-cellotriosyl-d-glucose (DP4). These oligosaccharides provided growth substrates for S. variabile and H. hathewayi with a preference for DP4 in the case of the latter species. There was increased transcription of a B. ovatus mixed-linkage-β-glucan utilization locus, as well as carbohydrate transporters in S. variabile and H. hathewayi when in batch coculture. Increased transcription of the β-glucan utilization locus did not occur in continuous culture. Evidence for interactions relating to provision of cobalamin, alterations to signaling, and modulation of the "stringent response" (an adaptation to nutrient deprivation) were detected. Overall, we established a bacterial consortium based on barley β-glucan in vitro, which can be used to investigate aspects of the functional blueprint of the human gut microbiota.IMPORTANCE The microbial community, mostly composed of bacterial species, residing in the human gut degrades and ferments polysaccharides derived from plants (dietary fiber) that would not otherwise be digested. In this way, the collective metabolic actions of community members extract additional energy from the human diet. While the variety of bacteria present in the microbial community is well known, the formation of bacterial consortia, and the consequent interactions that result in the digestion of dietary polysaccharides, has not been studied extensively. The importance of our work was the establishment, under laboratory conditions, of a consortium of gut bacteria that formed around a dietary constituent commonly present in cereals. This enabled the metabolic interplay between the bacterial species to be studied. This kind of knowledge is required to construct an interactive, metabolic blueprint of the microbial community that inhabits the human gut.

Keywords: RNAseq; bacterial consortium; beta-glucan; gut microbiota; whole-transcriptome analysis.

FIG 1

Growth of B. ovatus ATCC 8483^T, H. hathewayi DSM 13479^T, and S. variabile DSM 15176^T in pure batch cultures during anaerobic incubation at 37°C. Mean (standard error of the mean [SEM]) optical densities at A₆₀₀ of triplicate cultures are shown. (A) B. ovatus ATCC 8483^T growth in barley β-glucan (BG) medium compared to basal medium. (B) H. hathewayi DSM 13479^T growth in oligosaccharide (oligo) medium compared to basal medium. Note the diauxic growth profile in oligosaccharide medium where initial growth relied on constituents in basal medium, followed by utilization of oligosaccharides. (C) S. variabile DSM 15176^T growth in oligosaccharide (oligo) medium compared to basal medium.

FIG 2

Depolymerization of barley β-glucan by B. ovatus ATCC 8483^T in pure batch culture at 37°C. Size exclusion chromatography (SEC) profiles of uninoculated barley β-glucan medium, barley β-glucan standard, and supernatant from B. ovatus ATCC 8483^T culture in barley β-glucan medium incubated for 24 h.

FIG 3

High-performance anion-exchange chromatography (HPAEC) profiles of supernatants collected during the growth of B. ovatus ATCC 8483^T, S. variabile DSM 15176^T, and H. hathewayi DSM 13479^T in pure culture during anaerobic incubation at 37°C. Peaks corresponding to DP2 (probably laminaribiose), DP3 (3-O-β-cellobiosyl-d-glucose), and DP4 (3-O-β-cellotriosyl-d-glucose) oligosaccharides are indicated by reference to standards. Sampling times are indicated, and chromatographic profiles of samples that were analyzed with whole-transcriptomics analysis (RNA-seq) are colored red/orange. (A) B. ovatus ATCC 8483^T supernatants from barley β-glucan medium. (B) H. hathewayi DSM 13479^T supernatants from oligosaccharide medium. (C) S. variabile DSM 15176^T supernatants from oligosaccharide medium.

FIG 4

Populations of B. ovatus ATCC 8483^T, S. variabile DSM 15176^T, and H. hathewayi DSM 13479^T in batch (24 h of incubation) or continuous coculture and utilization of DP3 and DP4 oligosaccharides in continuous coculture. (A) CFU/ml of B. ovatus ATCC 8483^T, S. variabile DSM 15176^T, and H. hathewayi DSM 13479^T in combinations of batch coculture. Means (SEMs) of duplicate cultures are shown. (B) CFU/ml of B. ovatus ATCC 8483^T, S. variabile DSM 15176^T, and H. hathewayi DSM 13479^T in continuous coculture. Means (SEMs) of three chemostat runs are shown. (C) High-performance anion-exchange chromatography (HPAEC) profiles of supernatants collected from continuous coculture of B. ovatus ATCC 8483^T, S. variabile DSM 15176^T, and H. hathewayi DSM 13479^T. Profiles from three chemostat runs are shown. BO, B. ovatus ATCC 8483T in pure culture; BO+SV+HH, the three species in coculture. Peaks corresponding to DP3 (3-O-β-cellobiosyl-d-glucose) and DP4 (3-O-β-cellotriosyl-d-glucose) oligosaccharides are indicated.