Quantitative assay to detect bacterial glycan-degrading enzyme activities in mouse and human fecal samples

Steimle, Alex; Grant, Erica T.; Desai, Mahesh S.

doi:10.1016/j.xpro.2021.100326

Cited by 11 publications

(10 citation statements)

References 3 publications

(4 reference statements)

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“…Frozen fecal samples were stored and processed according to an established protocol. 16 The activity of three host mucus targeting bacterial enzymes, β- N -acetylglucosaminidase, sulfatase, α-fucosidase, and two dietary plant fiber glycan-targeting bacterial enzymes, α-galactosidase, and β-glucosidase were investigated. Details about bacterial enzymes, their biological substrate and the chemical substrate used for activity detection can be found in Table 1 .…”

Section: Methodsmentioning

confidence: 99%

Deprivation of dietary fiber in specific-pathogen-free mice promotes susceptibility to the intestinal mucosal pathogen Citrobacter rodentium

et al. 2021

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The change of dietary habits in Western societies, including reduced consumption of fiber, is linked to alterations in gut microbial ecology. Nevertheless, mechanistic connections between diet-induced microbiota changes that affect colonization resistance and enteric pathogen susceptibility are still emerging. We sought to investigate how a diet devoid of soluble plant fibers impacts the structure and function of a conventional gut microbiota in specific-pathogen-free (SPF) mice and how such changes alter susceptibility to a rodent enteric pathogen. We show that absence of dietary fiber intake leads to shifts in the abundances of specific taxa, microbiome-mediated erosion of the colonic mucus barrier, a reduction of intestinal barrier-promoting short-chain fatty acids, and increases in markers of mucosal barrier integrity disruption. Importantly, our results highlight that these low-fiber diet-induced changes in the gut microbial ecology collectively contribute to a lethal colitis by the mucosal pathogen Citrobacter rodentium , which is used as a mouse model for enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC, respectively). Our study indicates that modern, low-fiber Western-style diets might make individuals more prone to infection by enteric pathogens via the disruption of mucosal barrier integrity by diet-driven changes in the gut microbiota, illustrating possible implications for EPEC and EHEC infections.

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Section: Methodsmentioning

confidence: 99%

Deprivation of dietary fiber in specific-pathogen-free mice promotes susceptibility to the intestinal mucosal pathogen Citrobacter rodentium

et al. 2021

Self Cite

View full text Add to dashboard Cite

show abstract

“…Frozen fecal samples were stored and processed according to an established protocol 16 . The activity of three host mucus targeting bacterial enzymes, β- N -acetylglucosaminidase, sulfatase, α-fucosidase, and two dietary plant fiber glycan-targeting bacterial enzymes, α-galactosidase, and β-glucosidase were investigated.…”

Section: Methodsmentioning

confidence: 99%

Deprivation of dietary fiber in specific-pathogen-free mice promotes susceptibility to the intestinal mucosal pathogen Citrobacter rodentium

Neumann

Steimle

Grant

et al. 2021

Preprint

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The change of dietary habits in Western societies, including reduced consumption of fiber, is linked to alterations in gut microbial ecology. Nevertheless, mechanistic connections between diet-induced microbiota changes that affect colonization resistance and enteric pathogen susceptibility are still emerging. We sought to investigate how a diet devoid of soluble plant fibers impacts the structure and function of a conventional gut microbiota in specific-pathogen-free (SPF) mice and how such changes alter susceptibility to a rodent enteric pathogen. We show that absence of dietary fiber intake leads to shifts in the abundances of specific taxa, microbiome-mediated erosion of the colonic mucus barrier, a reduction of intestinal barrier-promoting short-chain fatty acids, and increases in markers of mucosal barrier integrity disruption. Importantly, our results highlight that these low fiber diet-induced changes in the gut microbial ecology collectively contribute to a lethal colitis by the mucosal pathogen Citrobacter rodentium, which is used as a mouse model for enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC, respectively). Our study indicates that modern, low-fiber Western diets might make individuals more prone to infection by enteric pathogens via the disruption of mucosal barrier integrity by diet-driven changes in the gut microbiota, illustrating possible implications for EPEC and EHEC infections.

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“…Enzymatic activities of β-glucosidase, α-galactosidase, α-fucosidase, β-N-acetylgluco saminidase and sulfatase were detected from fecal samples stored at −20 • C as described previously [44]. In brief, bacterial glycan-degrading enzymes were solubilized from the fecal samples by incubation in a lysozyme-, DNase I-and Triton X-100-containing lysis buffer on ice followed by sonification and removal of unsolubilized material by centrifugation.…”

Section: Detection Of Glycan-degrading Enzyme Activities In Fecal Samplesmentioning

confidence: 99%

“…For detection of enzymatic activities, equal amounts of protein were incubated with enzyme-specific p-nitrophenol-coupled substrates, and the substrate turnover (pnitrophenol release) was monitored by kinetic measurements of optical density at 405 nm. For details on buffers, substrates and final computation of enzymatic activities from optical density data, refer to [44].…”

Section: Detection Of Glycan-degrading Enzyme Activities In Fecal Samplesmentioning

confidence: 99%

Concentrated Raw Fibers Enhance the Fiber-Degrading Capacity of a Synthetic Human Gut Microbiome

Steimle

Neumann

Grant

et al. 2021

IJMS

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The consumption of prebiotic fibers to modulate the human gut microbiome is a promising strategy to positively impact health. Nevertheless, given the compositional complexity of the microbiome and its inter-individual variances, generalized recommendations on the source or amount of fiber supplements remain vague. This problem is further compounded by availability of tractable in vitro and in vivo models to validate certain fibers. We employed a gnotobiotic mouse model containing a 14-member synthetic human gut microbiome (SM) in vivo, characterized a priori for their ability to metabolize a collection of fibers in vitro. This SM contains 14 different strains belonging to five distinct phyla. Since soluble purified fibers have been a common subject of studies, we specifically investigated the effects of dietary concentrated raw fibers (CRFs)—containing fibers from pea, oat, psyllium, wheat and apple—on the compositional and functional alterations in the SM. We demonstrate that, compared to a fiber-free diet, CRF supplementation increased the abundance of fiber-degraders, namely Eubacterium rectale, Roseburia intestinalis and Bacteroides ovatus and decreased the abundance of the mucin-degrader Akkermansia muciniphila. These results were corroborated by a general increase of bacterial fiber-degrading α-glucosidase enzyme activity. Overall, our results highlight the ability of CRFs to enhance the microbial fiber-degrading capacity.

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Quantitative assay to detect bacterial glycan-degrading enzyme activities in mouse and human fecal samples

Cited by 11 publications

References 3 publications

Deprivation of dietary fiber in specific-pathogen-free mice promotes susceptibility to the intestinal mucosal pathogen Citrobacter rodentium

Deprivation of dietary fiber in specific-pathogen-free mice promotes susceptibility to the intestinal mucosal pathogen Citrobacter rodentium

Deprivation of dietary fiber in specific-pathogen-free mice promotes susceptibility to the intestinal mucosal pathogen Citrobacter rodentium

Concentrated Raw Fibers Enhance the Fiber-Degrading Capacity of a Synthetic Human Gut Microbiome

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