乳酸菌分离培养基对粪便样品筛选能力的探究

Study on the screening capability of lactic acid bacteria isolation media for fecal samples

目的探究在不同氧气条件下,不同的常用乳酸菌分离培养基分离婴儿粪便样品中微生物的能力,找出最合适的益生菌分离培养基及相关分离条件。方法以婴儿粪便为原料,在利用乳酸杆菌选择性琼脂培养基(Lactobacillus selective broth,LBS)、MRS以及MRS5.23种培养基对同一样品在有氧和厌氧条件下分别进行涂布培养。随后对样本内菌群进行16S rRNA扩增子测序,通过对测序数据的菌落组成、α多样性、β多样性及后续代谢的模拟分析,阐释了不同培养基对粪便样品的筛选特征,包括筛选后菌群的组内多样性、组间多样性、菌群功能预测以及培养基中乳酸菌所占比例等。结果对于婴儿粪便样品,所测5种分离条件均能对肠道中的乳酸杆菌进行有效分离。在厌氧条件下除LBS培养基外,各培养基中均有双歧杆菌出现,所占比率约在20%~30%之间,同时有氧环境下的MRS培养基中也存在少量双歧杆菌,所占比率约为5%。结论对于粪便样品中乳杆菌分离能力最适的条件是37℃下厌氧条件的LBS培养基,对双歧杆菌分离能力最适的条件是37℃下厌氧环境下的MRS培养基。

Objective To explore the ability of different commonly used lactic acid bacteria isolation media to separate microorganisms from infant feces samples under different oxygen conditions,and to find the most suitable probiotic isolation media and related isolation conditions.Methods Taking infant feces as raw materials,the sample was spread and cultured by using three types of media including Lactobacillus selective broth(LBS),MRS and MRS5.2 media under aerobic and anaerobic conditions.Subsequently,16 S rRNA amplicon sequencing was performed on the bacterial population in the sample.Through the analysis of the colony composition,αdiversity,β-diversity and simulated metabolism of the sequenced data,the screening characteristics of different media for fecal sample were explained,including the intra-group diversity,the inter-group diversity of the selected flora,the prediction of flora function,and the proportion of lactic acid bacteria in the tested culture conditions.Results For infant feces samples,all the five separation conditions can effectively separate Lactobacillus from the intestine.Under anaerobic conditions,Bifidobacterium were found in all media except LBS media,which accounted for 20%-30%.At the same time,a small amount of Bifidobacterium was also found in MRS medium under aerobic conditions,accounting for about 5%.Conclusion For infant fecal samples,LBS media at 37°C under anaerobic conditions was the optimal condition for Lactobacillus isolation,and MRS medium at 37°C under anaerobic conditions was the best for Bifidobacterium isolation.