接种固定化粪菌和培养基种类对体外结肠菌群发酵的影响

Effect of microbiota immobilization and medium type on mimic continuous colon fermentation in vitro

目的研究三种模拟结肠发酵培养基和粪菌固定化对体外结肠发酵体系菌群构成的影响,为建立高度模拟结肠体外结肠发酵提供参考。方法采集健康女性成人粪便,粪便制备悬液或用结冷胶-黄原胶固定化粪菌接种于模拟结肠发酵反应器,分别用三种培养基按0.07 m L/h稀释率进行连续发酵10 d,用末端限制性片段长度多态性(T-RFLP)与高通量测序技术分析菌群多样性,并分析发酵液短链脂肪酸含量。结果 T-RFLP分析显示,培养基类型主要影响粪菌固定体系菌群α多样性达到稳定的时间和多样性。β多样性分析显示,第10天时培养基Ⅲ悬液培养和培养基I固定化培养系统的菌群构成与粪便菌群最相近。而培养基Ⅲ在第10天达到较高而稳定的丁酸浓度。结论由于比粪菌固定培养更易操作,培养基Ⅲ悬液培养适于模拟人结肠发酵。

Objective To study the effects of three kinds of medium and faecal immobilization on microbiota composition in proximal colonic fermentation in vitro, and provide scientific support for the establishment of fermentation system highly mimicking proximal colon. Methods Feces from a healthy adult woman were made into 10% （w/v） slurries, or immobilized using gellan gum-xanthan gum, then inoculated into the reac- tor mimicking colon fermentation. Three media were tested for 10 days of continuous fermentation respec tively with dilution rate of 0.07 mL/h. Terminal restriction fragment length polymorphism （T RFLP） and high-throughput sequencing were used to analyze bacterial diversity. Short chain fatty acids were analyzed by using HPLC. Results T-RFLP analysis showed that different media affected the time to stability of commu- nity and alpha-diversity. According to beta diversity analysis, the bacterial composition in medium III with out immobilization and medium I with immobilization were more similar to that of feces at 10th day. Medium III reached a high and stable concentration of butyric acid on the 10th day. Conclusion Taking operability and effect into consideration, medium III without fecal immobilization is suitable for simulating human colon fermentation.