Novel Polyfermentor Intestinal Model (PolyFermS) for Controlled Ecological Studies: Validation and Effect of pH

In vitro gut fermentation modeling offers a useful platform for ecological studies of the intestinal microbiota. In this study we describe a novel Polyfermentor Intestinal Model (PolyFermS) designed to compare the effects of different treatments on the same complex gut microbiota. The model operated in conditions of the proximal colon is composed of a first reactor containing fecal microbiota immobilized in gel beads, and used to continuously inoculate a set of parallel second-stage reactors. The PolyFermS model was validated with three independent intestinal fermentations conducted for 38 days with immobilized human fecal microbiota obtained from three child donors. The microbial diversity of reactor effluents was compared to donor feces using the HITChip, a high-density phylogenetic microarray targeting small subunit rRNA sequences of over 1100 phylotypes of the human gastrointestinal tract. Furthermore, the metabolic response to a decrease of pH from 5.7 to 5.5, applied to balance the high fermentative activity in inoculum reactors, was studied. We observed a reproducible development of stable intestinal communities representing major taxonomic bacterial groups at ratios similar to these in feces of healthy donors, a high similarity of microbiota composition produced in second-stage reactors within a model, and a high time stability of microbiota composition and metabolic activity over 38 day culture. For all tested models, the pH-drop of 0.2 units in inoculum reactors enhanced butyrate production at the expense of acetate, but was accompanied by a donor-specific reorganization of the reactor community, suggesting a concerted metabolic adaptation and trigger of community-specific lactate or acetate cross-feeding pathways in response to varying pH. Our data showed that the PolyFermS model allows the stable cultivation of complex intestinal microbiota akin to the fecal donor and can be developed for the direct comparison of different experimental conditions in parallel reactors continuously inoculated with the exact same microbiota.

体外肠道发酵模型为肠道菌群的生态学研究提供了实用研究平台。本研究介绍一种新型发酵罐肠道模型（Polyfermentor Intestinal Model，PolyFermS），其设计目的是比较不同处理对同一复杂肠道菌群的影响。该模型模拟近端结肠环境，由搭载固定于凝胶微球中粪便菌群的一级反应器，以及用于持续接收该反应器接种物的多组平行二级反应器组成。本研究以3名儿童供者的固定化人类粪便菌群为接种物，开展了3组独立的肠道发酵实验，持续运行38天以验证PolyFermS模型的有效性。研究采用HITChip——一种靶向人类胃肠道1100余种系统型小亚基核糖体RNA（rRNA）序列的高密度系统发育微阵列，将反应器流出物的微生物多样性与供者粪便进行比对。此外，本研究还探究了为平衡接种物反应器的高发酵活性，将其pH从5.7降至5.5所引发的代谢响应。研究结果显示，该模型可稳定形成涵盖主要细菌分类类群的肠道菌群群落，其组成比例与健康供者粪便菌群相似；同一模型内二级反应器产出的菌群组成具有高度相似性，且培养38天期间，菌群组成与代谢活性均表现出极高的时间稳定性。在所有测试模型中，接种物反应器0.2单位的pH降幅均提升了丁酸产量，但以乙酸产量为代价；同时伴随反应器菌群组成发生供者特异性重构，这表明菌群会协同发生代谢适应性变化，并启动群落特异性的乳酸或乙酸交叉喂养通路以响应pH变化。本研究数据表明，PolyFermS模型可稳定培养与供者粪便菌群相似的复杂肠道菌群，且可用于在持续接种完全一致菌群的平行反应器中，直接比较不同实验处理的效果。