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ARS Home » Northeast Area » Wyndmoor, Pennsylvania » Eastern Regional Research Center » Dairy and Functional Foods Research » Research » Research Project #433215

Research Project: In Vitro Human Intestinal Microbial Ecosystem: Effects of Diet

Location: Dairy and Functional Foods Research

Project Number: 8072-41000-102-00-D
Project Type: In-House Appropriated

Start Date: Aug 29, 2017
End Date: May 1, 2020

Objective:
1: Establish a gut microbiota community utilizing the twin-Simulator of the Human Intestinal Microbial Ecology (TWINSHIME®) system. 2: Evaluate changes in the bacterial a) community composition b) metabolome, and c) proteome in response to dietary components. 2a: Evaluate changes in the population composition, metabolome, and proteome of the gut microbiota using the TWINSHIME® in response to fluctuations in the pH of the large intestinal regions. 2b: Evaluate alterations in the population composition, metabolome, and proteome of the gut microbiota using the TWINSHIME® in response to defined dietary interventions. In particular fat-free or full-fat milk and whole wheat or refined wheat flour.

Approach:
This project will study the effects of dietary components on the human gut microbiota of the large intestine. This will be done using the TWINSHIME® system, which is a dynamic, in vitro system capable of simulating the physiological conditions of the human gastrointestinal tract. This system is comprised of two sets of bioreactors (SHIME 1 and 2) arranged in parallel to mimic the stomach, small intestine, and the large intestine, which is divided into the ascending, transverse, and descending regions. In the first objective, the ability for the TWINSHIME® system to establish a stable human gut microbiota community, representative of the distinct regions of the large intestine will be examined. Data from Next Generation DNA sequencing and short chain fatty acids (SCFA) analysis will be used to confirm stability and demonstrate that once it is achieved, it remains the same until the experiment is terminated. Furthermore, the proteomics and metabolomics research will enable establishment of the correlation of the change in composition with specifically functional expression of gut microbiota. In the second objective, changes in the bacterial a) community composition b) metabolome, and c) proteome, in response to alterations of pH and the dietary components milk and whole wheat will be measured. To analyze the response of the microbiota to changes in pH, the pH of the colon reactors in SHIME 1 will be lowered, while the pH of the colon reactors in SHIME 2 will be increased. In order to determine changes to the bacterial community, metabolome, and proteome in response to milk, SHIME 1 will be supplemented with 8 ounces of fat-free milk per feeding and SHIME 2 supplemented with 8 ounces of full-fat milk per feeding. In order to test the effect of wheat flour on the gut microbiota population and/or metabolome, SHIME 1 will be supplemented with refined wheat flour and SHIME 2 will be supplemented with whole wheat flour. During these experiments, samples will be harvested every three days and subjected to analysis. Data from 16S rRNA sequencing will be used to determine community composition; Gas and Liquid chromatography, coupled with mass spectrometry, and a MALDI-TOF/TOF-MS/MS will be used to determine changes in the metabolome (SCFA, amino acids, volatiles, peptides, sugars, and lipids) and the proteome. By compiling these results, the effects of pH change, milk, and wheat on the community composition, metabolome and proteome of the gut microbiota of the individual colon regions can be accurately determined.