Inducible CRISPR-targeted "knockdown" of human gut Bacteroides in gnotobiotic mice discloses glycan utilization strategies.

Understanding how members of the human gut microbiota prioritize nutrient resources is one component of a larger effort to decipher the mechanisms defining microbial community robustness and resiliency in health and disease. This knowledge is foundational for development of microbiota-directed therapeutics. To model how bacteria prioritize glycans in the gut, germfree mice were colonized with 13 human gut bacterial strains, including seven saccharolytic Bacteroidaceae species. Animals were fed a Western diet supplemented with pea fiber. After community assembly, an inducible CRISPR-based system was used to selectively and temporarily reduce the absolute abundance of Bacteroides thetaiotaomicron or B. cellulosilyticus by 10- to 60-fold. Each knockdown resulted in specific, reproducible increases in the abundances of other Bacteroidaceae and dynamic alterations in their expression of genes involved in glycan utilization. Emergence of these "alternate consumers" was associated with preservation of community saccharolytic activity. Using an inducible system for CRISPR base editing in vitro, we disrupted translation of transporters critical for utilizing dietary polysaccharides in Phocaeicola vulgatus, a B. cellulosilyticus knockdown-responsive taxon. In vitro and in vivo tests of the resulting P. vulgatus mutants allowed us to further characterize mechanisms associated with its increased fitness after knockdown. In principle, the approach described can be applied to study utilization of a range of nutrients and to preclinical efforts designed to develop therapeutic strategies for precision manipulation of microbial communities.

Bifidobacterium infantis treatment promotes weight gain in Bangladeshi infants with severe acute malnutrition.

Disrupted development of the gut microbiota is a contributing cause of childhood malnutrition. Bifidobacterium longum subspecies infantis is a prominent early colonizer of the infant gut that consumes human milk oligosaccharides (HMOs). We found that the absolute abundance of Bifidobacterium infantis is lower in 3- to 24-month-old Bangladeshi infants with severe acute malnutrition (SAM) compared to their healthy age-matched counterparts. A single-blind, placebo-controlled trial (SYNERGIE) was conducted in 2- to 6-month-old Bangladeshi infants with SAM. A commercial U.S. donor-derived B. infantis strain (EVC001) was administered daily with or without the HMO lacto-N-neotetraose for 28 days. This intervention increased fecal B. infantis abundance in infants with SAM, although to levels still 10- to 100-fold lower than in untreated healthy controls. EVC001 treatment promoted weight gain that was associated with reduced intestinal inflammation markers in infants with SAM. We cultured fecal B. infantis strains from Bangladeshi infants and colonized gnotobiotic mice with these cultured strains. The gnotobiotic mice were fed a diet representative of that consumed by 6-month-old Bangladeshi infants, with or without HMO supplementation. One B. infantis strain, Bg_2D9, expressing two gene clusters involved in uptake and utilization of N-glycans and plant-derived polysaccharides, exhibited superior fitness over EVC001. The fitness advantage of Bg_2D9 was confirmed in a gnotobiotic mouse model of mother-to-infant gut microbiota transmission where dams received a pretreatment fecal community from a SAM infant in the SYNERGIE trial. Whether Bg_2D9 is superior to EVC001 for treating malnourished infants who consume a diet with limited breastmilk requires further clinical testing.

Combined Prebiotic and Microbial Intervention Improves Oral Cholera Vaccination Responses in a Mouse Model of Childhood Undernutrition.

Undernourished children in low-income countries often exhibit poor responses to oral vaccination. Perturbed microbiota development is linked to undernutrition, but whether and how microbiota changes affect vaccine responsiveness remains unclear. Here, we show that gnotobiotic mice colonized with microbiota from undernourished Bangladeshi children and fed a Bangladeshi diet exhibited microbiota-dependent differences in mucosal IgA responses to oral vaccination with cholera toxin (CT). Supplementation with a nutraceutical consisting of spirulina, amaranth, flaxseed, and micronutrients augmented CT-IgA production. Mice initially colonized with a microbiota associated with poor CT responses exhibited improved immunogenicity upon invasion of bacterial taxa from cagemates colonized with a more "responsive" microbiota. Additionally, a consortium of five cultured bacterial invaders conferred augmented CT-IgA responses in mice fed the supplemented diet and colonized with the "hypo-responsive" community. These results provide preclinical proof-of-concept that diet and microbiota influence mucosal immune responses to CT vaccination and identify a candidate synbiotic formulation.