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      Procedures for Fecal Microbiota Transplantation in Murine Microbiome Studies


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          Fecal microbiota transplantation (FMT) has been widely recognized as an approach to determine the microbiome’s causal role in gut dysbiosis-related disease models and as a novel disease-modifying therapy. Despite potential beneficial FMT results in various disease models, there is a variation and complexity in procedural agreement among research groups for performing FMT. The viability of the microbiome in feces and its successful transfer depends on various aspects of donors, recipients, and lab settings. This review focuses on the technical practices of FMT in animal studies. We first document crucial factors required for collecting, handling, and processing donor fecal microbiota for FMT. Then, we detail the description of gut microbiota depletion methods, FMT dosages, and routes of FMT administrations in recipients. In the end, we describe assessments of success rates of FMT with sustainability. It is critical to work under the anaerobic condition to preserve as much of the viability of bacteria. Utilization of germ- free mice or depletion of recipient gut microbiota by antibiotics or polyethylene glycol are two common recipient preparation approaches to achieve better engraftment. Oral-gastric gavage preferred by most researchers for fast and effective administration of FMT in mice. Overall, this review highlights various methods that may lead to developing the standard and reproducible protocol for FMT.

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          Structure, Function and Diversity of the Healthy Human Microbiome

          Studies of the human microbiome have revealed that even healthy individuals differ remarkably in the microbes that occupy habitats such as the gut, skin, and vagina. Much of this diversity remains unexplained, although diet, environment, host genetics, and early microbial exposure have all been implicated. Accordingly, to characterize the ecology of human-associated microbial communities, the Human Microbiome Project has analyzed the largest cohort and set of distinct, clinically relevant body habitats to date. We found the diversity and abundance of each habitat’s signature microbes to vary widely even among healthy subjects, with strong niche specialization both within and among individuals. The project encountered an estimated 81–99% of the genera, enzyme families, and community configurations occupied by the healthy Western microbiome. Metagenomic carriage of metabolic pathways was stable among individuals despite variation in community structure, and ethnic/racial background proved to be one of the strongest associations of both pathways and microbes with clinical metadata. These results thus delineate the range of structural and functional configurations normal in the microbial communities of a healthy population, enabling future characterization of the epidemiology, ecology, and translational applications of the human microbiome.
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            The microbial metabolites, short-chain fatty acids, regulate colonic Treg cell homeostasis.

            Regulatory T cells (Tregs) that express the transcription factor Foxp3 are critical for regulating intestinal inflammation. Candidate microbe approaches have identified bacterial species and strain-specific molecules that can affect intestinal immune responses, including species that modulate Treg responses. Because neither all humans nor mice harbor the same bacterial strains, we posited that more prevalent factors exist that regulate the number and function of colonic Tregs. We determined that short-chain fatty acids, gut microbiota-derived bacterial fermentation products, regulate the size and function of the colonic Treg pool and protect against colitis in a Ffar2-dependent manner in mice. Our study reveals that a class of abundant microbial metabolites underlies adaptive immune microbiota coadaptation and promotes colonic homeostasis and health.
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              Metabolites produced by commensal bacteria promote peripheral regulatory T cell generation

              Intestinal microbes provide multicellular hosts with nutrients and confer resistance to infection. The delicate balance between pro- and anti-inflammatory mechanisms, essential for gut immune homeostasis, is affected by the composition of the commensal microbial community. Regulatory T (Treg) cells expressing transcription factor Foxp3 play a key role in limiting inflammatory responses in the intestine 1 . Although specific members of the commensal microbial community have been found to potentiate the generation of anti-inflammatory Treg or pro-inflammatory Th17 cells 2-6 , the molecular cues driving this process remain elusive. Considering the vital metabolic function afforded by commensal microorganisms, we hypothesized that their metabolic by-products are sensed by cells of the immune system and affect the balance between pro- and anti-inflammatory cells. We found that a short-chain fatty acid (SCFA), butyrate, produced by commensal microorganisms during starch fermentation, facilitated extrathymic generation of Treg cells. A boost in Treg cell numbers upon provision of butyrate was due to potentiation of extrathymic differentiation of Treg cells as the observed phenomenon was dependent upon intronic enhancer CNS1, essential for extrathymic, but dispensable for thymic Treg cell differentiation 1, 7 . In addition to butyrate, de novo Treg cell generation in the periphery was potentiated by propionate, another SCFA of microbial origin capable of HDAC inhibition, but not acetate, lacking this activity. Our results suggest that bacterial metabolites mediate communication between the commensal microbiota and the immune system, affecting the balance between pro- and anti-inflammatory mechanisms.

                Author and article information

                Front Cell Infect Microbiol
                Front Cell Infect Microbiol
                Front. Cell. Infect. Microbiol.
                Frontiers in Cellular and Infection Microbiology
                Frontiers Media S.A.
                21 September 2021
                : 11
                : 711055
                [1]Department of Medicine, University of Connecticut (UConn) Health , Farmington, CT, United States
                Author notes

                Edited by: Felix Broecker, Idorsia Pharmaceuticals Ltd, Switzerland

                Reviewed by: Yuanqiang Zou, Beijing Genomics Institute (BGI), China; Jane Adair Mullaney, AgResearch Ltd, New Zealand

                *Correspondence: Yanjiao Zhou, yazhou@ 123456uchc.edu

                This article was submitted to Microbiome in Health and Disease, a section of the journal Frontiers in Cellular and Infection Microbiology

                Copyright © 2021 Bokoliya, Dorsett, Panier and Zhou

                This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

                : 18 May 2021
                : 24 August 2021
                Page count
                Figures: 1, Tables: 2, Equations: 0, References: 165, Pages: 14, Words: 7600
                Funded by: National Institute of Neurological Disorders and Stroke , doi 10.13039/100000065;
                Cellular and Infection Microbiology

                Infectious disease & Microbiology
                fecal microbiota transplantation,mice,microbiome,engraftment,procedure


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