Table_2_Use of Changestat for Growth Rate Studies of Gut Microbiota.XLSX

Human colon microbiota, composed of hundreds of different species, is closely associated with several health conditions. Controlled in vitro cultivation and up-to-date analytical methods make possible the systematic evaluation of the underlying mechanisms of complex interactions between the members of microbial consortia. Information on reproducing fecal microbial consortia can be used for various clinical and biotechnological applications. In this study, chemostat and changestat cultures were used to elucidate the effects of the physiologically relevant range of dilution rates on the growth and metabolism of adult fecal microbiota. The dilution rate was kept either at D = 0.05 or D = 0.2 1/h in chemostat cultures, while gradually changing from 0.05 to 0.2 1/h in the A-stat and from 0.2 to 0.05 1/h in the De-stat. Apple pectin as a substrate was used in the chemostat experiments and apple pectin or birch xylan in the changestat experiments, in the presence of porcine mucin in all cases. The analyses were comprised of HPLC for organic acids, UPLC for amino acids, GC for gas composition, 16S-rDNA sequencing for microbial composition, and growth parameter calculations. It was shown that the abundance of most bacterial taxa was determined by the dilution rate on both substrates. Bacteroides ovatus, Bacteroides vulgatus, and Faecalibacterium were prevalent within the whole range of dilution rates. Akkermansia muciniphila and Ruminococcaceae UCG-013 were significantly enriched at D = 0.05 1/h, while Bacteroides caccae, Lachnospiraceae unclassified and Escherichia coli clearly preferred D = 0.2 1/h. In the chemostat cultures, the production of organic acids and gases from pectin was related to the dilution rate. The ratio of acetate, propionate and butyrate was 5:2:1 (D = 0.05 1/h) and 14:2:1 (D = 0.2 1/h). It was shown that the growth rate-related characteristics of the fecal microbiota were concise in both directions between D = 0.05 and 0.2 1/h. Reproducible adaptation of the fecal microbiota was shown in the continuous culture with a changing dilution rate: changestat. Consortia cultivation is a promising approach for research purposes and several biotechnological applications, including the production of multi-strain probiotics and fecal transplantation mixtures.

人类结肠微生物群，由数百种不同的物种组成，与多种健康状态密切相关。通过精确的体外培养及先进的分析技术，使得对微生物群落成员之间复杂交互作用的潜在机制进行系统性评估成为可能。关于复制粪便微生物群的信息，可用于多种临床及生物技术应用。在本研究中，通过使用化学稳态和变化稳态培养，旨在阐明与生理相关范围内的稀释速率对成人粪便微生物群生长和代谢的影响。在化学稳态培养中，稀释速率保持在D = 0.05或D = 0.2 1/h，而在A-stat和De-stat中，稀释速率则从0.05逐渐变化至0.2 1/h，以及从0.2变化至0.05 1/h。所有实验中均使用了猪粘蛋白，在化学稳态实验中使用苹果果胶作为底物，在变化稳态实验中使用苹果果胶或桦木木聚糖，所有实验均在猪粘蛋白存在的情况下进行。分析包括有机酸的高效液相色谱分析、氨基酸的液相色谱-质谱联用分析、气体组成的气相色谱分析、16S-rDNA测序分析微生物组成，以及生长参数的计算。研究表明，在两种底物上，大多数细菌类群的丰度均由稀释速率决定。Bacteroides ovatus、Bacteroides vulgatus和Faecalibacterium在所有稀释速率范围内普遍存在。Akkermansia muciniphila和Ruminococcaceae UCG-013在D = 0.05 1/h时显著富集，而Bacteroides caccae、Lachnospiraceae未分类群和Escherichia coli则明显偏好D = 0.2 1/h。在化学稳态培养中，从果胶中产生的有机酸和气体的产量与稀释速率相关。乙酸、丙酸和丁酸的比例为5:2:1（D = 0.05 1/h）和14:2:1（D = 0.2 1/h）。研究表明，粪便微生物群的生长速率相关特征在D = 0.05和0.2 1/h之间呈现双向简洁关系。在连续培养中，具有变化稀释速率的changestat展示了粪便微生物群的可重复适应性。群落培养作为一种有前景的研究方法，以及在多菌株益生菌和粪便移植混合物的生产等多种生物技术应用中，均显示出其广阔的应用前景。