A mutant fitness compendium in Bifidobacteria reveals molecular determinants of colonization and host-microbe interactions

Abstract

Bifidobacteria commonly represent a dominant constituent of human gut microbiomes during infancy, influencing nutrition, immune development, and resistance to infection. Despite interest as a probiotic therapy, predicting the nutritional requirements and health-promoting effects of Bifidobacteria is challenging due to major knowledge gaps. To overcome these deficiencies, we used large-scale genetics to create a compendium of mutant fitness inBifidobacterium breve(Bb). We generated a high density, randomly barcoded transposon insertion pool inBb, and used this pool to determineBbfitness requirements during colonization of germ-free mice and chickens with multiple diets and in response to hundreds ofin vitroperturbations. To enable mechanistic investigation, we constructed an ordered collection of insertion strains covering 1462 genes. We leveraged these tools to improve models of metabolic pathways, reveal unexpected host- and diet-specific requirements for colonization, and connect the production of immunomodulatory molecules to growth benefits. These resources will greatly reduce the barrier to future investigations of this important beneficial microbe.

Unveiling metabolic pathways of selected plant-derived glycans by Bifidobacterium pseudocatenulatum

“…In order to validate the predicted starch-degrading activity of the product of amy1 , as its activity was not obvious from the previous enzymatic assays, the gene was cloned under the control of a constitutive promoter (see Materials and Methods), resulting in plasmid pBC1.2: amy1 . Plasmid pBC1.2: amy1 was then introduced in B. breve UCC2003:ΔApuBP8, which is a derivative of UCC2003 in which the apuB gene had been interrupted by transposon mutagenesis, thus making this strain unable to grow in starch or starch-like glycans ( O’Connell Motherway et al, 2008 ; Shiver et al, 2021 , 2023 ).…”

Systematic mapping of antibiotic cross-resistance and collateral sensitivity with chemical genetics

“…As proof-of-principle we focused on published chemical genetics data from E. coli 31 , because of the large number of antibiotics screened at different concentrations and the extensive benchmarking. In the future, similar analyses can be expanded to other available datasets in the same or other species 34,47,[53][54][55][56] . Such datasets will inevitably increase with time, as genome-wide mutant libraries are becoming available in tens of species and even more strains 57,58 , whether those are arrayed or pooled 29,59 , and constructed by targeted deletions [60][61][62] , transposon insertions 59,63 , or CRISPRi knockdowns 53,64 .…”

Integrative genomic reconstruction of carbohydrate utilization networks in bifidobacteria: global trends, local variability, and dietary adaptation

“…For example, the end products of bifidobacterial carbohydrate metabolism, lactate and acetate, can inhibit pathogen colonization [19][20][21] and serve as a basis for cross-feeding with various community members 22,23 . Additionally, multiple Bifidobacterium species produce aromatic lactic acids and other metabolites that modulate the immune system [24][25][26][27] . Given these beneficial properties, some bifidobacterial strains are commonly administered as probiotics [28][29][30][31][32] .…”