Establishing a mucosal gut microbial community in vitro using an artificial simulator

The Twin Simulator of the Human Intestinal Microbial Ecosystem (TWINSHIME®) was initially developed to study the luminal gut microbiota of the ascending (AC), transverse (TC), and descending (DC) colon regions. Given the unique composition and potential importance of the mucosal microbiota for human health, the TWINSHIME was recently adapted to simulate the mucosal microbiota as well as the luminal community. It has been previously demonstrated that the luminal community in the TWINSHIME reaches a steady state within two weeks post inoculation, and is able to differentiate into region specific communities. However, less is known regarding the mucosal community structure and dynamics. During the current study, the luminal and mucosal communities in each region of the TWINSHIME were evaluated over the course of six weeks. Based on 16S rRNA gene sequencing and short chain fatty acid analysis, it was determined that both the luminal and mucosal communities reached stability 10–20 days after inoculation, and remained stable until the end of the experiment. Bioinformatics analysis revealed the formation of unique community structures between the mucosal and luminal phases in all three colon regions, yet these communities were similar to the inoculum. Specific colonizers of the mucus mainly belonged to the Firmicutes phylum and included Lachnospiraceae (AC/TC/DC), Ruminococcaceae and Eubacteriaceae (AC), Lactobacillaceae (AC/TC), Clostridiaceae and Erysipelotrichaceae (TC/DC). In contrast, Bacteroidaceae were enriched in the gut lumen of all three colon regions. The unique profile of short chain fatty acid (SCFA) production further demonstrated system stability, but also proved to be an area of marked differences between the in vitro system and in vivo reports. Results of this study demonstrate that it is possible to replicate the community structure and composition of the gut microbiota in vitro. Through implementation of this system, the human gut microbiota can be studied in a dynamic and continuous fashion.人类肠道微生物组双模拟系统（Twin Simulator of the Human Intestinal Microbial Ecosystem，商标为TWINSHIME®）最初专为研究升结肠（ascending colon, AC）、横结肠（transverse colon, TC）及降结肠（descending colon, DC）的肠腔菌群而开发。鉴于黏膜菌群独特的组成及其对人类健康的潜在重要性，研究团队近期对TWINSHIME®进行了改造，使其可同时模拟黏膜菌群与肠腔菌群群落。既往研究已证实，TWINSHIME®的肠腔菌群可在接种后两周内达到稳态，并可分化为区域特异性菌群。然而，目前对于黏膜菌群的群落结构与动态变化的认知仍较为匮乏。本研究中，研究人员对TWINSHIME®各结肠区域的肠腔与黏膜菌群开展了为期六周的跟踪评估。基于16S rRNA基因测序与短链脂肪酸（short chain fatty acid, SCFA）分析结果，研究证实肠腔与黏膜菌群均在接种后10~20天达到稳定状态，并持续稳定至实验结束。生物信息学分析显示，三个结肠区域的黏膜菌群与肠腔菌群均形成了独特的群落结构，但二者均与接种起始菌群具有相似性。黏液层的特异性定植菌主要隶属于厚壁菌门（Firmicutes），包括毛螺菌科（Lachnospiraceae，AC/TC/DC）、瘤胃球菌科（Ruminococcaceae）与真杆菌科（Eubacteriaceae，仅见于AC）、乳杆菌科（Lactobacillaceae，AC/TC）、梭菌科（Clostridiaceae）与红蝽杆菌科（Erysipelotrichaceae，TC/DC）。与之相对，拟杆菌科（Bacteroidaceae）在三个结肠区域的肠腔中均呈现富集态势。短链脂肪酸的独特产生谱进一步验证了该系统的稳定性，同时也揭示了该体外系统与体内研究结果之间存在显著差异。本研究结果表明，体外模拟肠道菌群的群落结构与组成具备可行性。通过应用该系统，可实现对人类肠道菌群的动态、连续式研究。