初治结核病患者肠道菌群变性梯度凝胶电泳指纹图谱分析

doi:10.3969/j.issn.1000-6621.2019.09.013

摘要/Abstract

摘要：

目的分析正常人群与初治结核病患者肠道菌群的差异。方法于2017年8月至2018年7月每月固定时间在西电集团医院收集当天符合入组标准的健康体检成年人粪便标本2~3份,收集30份标本后停止纳入,作为对照组;收集2017年8月至2018年7月西安市胸科医院诊断为初治结核病患者的粪便标本,收集30份标本后停止纳入,作为结核组。利用变性梯度凝胶电泳技术分析对照组与结核组肠道菌群的多样性、相似性,以及变性梯度凝胶电泳主要条带测序后各组菌群所占比例。结果对照组的条带数[(13.57±3.37)个]和菌群多样性指数(H'指数;2.55±0.27)均明显高于结核组[(8.60±4.19)个和1.99±0.52],差异均有统计学意义(t=12.55,P<0.001;t=6.75,P<0.001)。组内Dice 相似性系数(Cs)的比较显示,对照组与结核组的中位数(四分位数)[22.90%(16.20%,29.30%)和35.35%(23.73%,44.98%)]间的差异有统计学意义(Z=-73.38,P<0.001)。测序比对结果显示,对照组和结核组的主要肠道菌群均以Bacteroides属为主,分别占73.96%(179/242)和79.40%(158/199)。但结核组中Prevotella菌属构成比(9.04%,18/199)较对照组(24.38%,59/242)有所下降,差异有统计学意义(χ ²=17.82,P<0.001),结核组Uncultured bacterium菌属的构成比(11.56%,23/199)高于对照组(0,0/242),差异有统计学意义(χ ²=29.51,P<0.001)。结论健康人群与结核病患者肠道菌群因机体免疫功能的不同存在差异,结核病的发生可能会造成Prevotella菌属和Uncultured bacterium菌属构成出现变化。

关键词: 结核, 肠, 细菌, 聚脱氧核糖核酸类, 变性梯度凝胶电泳

Abstract:

Objective To analyze the difference in gut microbiota between normal and newly diagnosed tuberculosis patients.Methods From August 2017 to July 2018, 2 to 3 copies of the fecal specimens of healthy medical examinations that met the inclusion criteria were collected at the fixed time every month in the Xi’dian Group Hospital. Specimens collection was stopped after collecting 30 specimens which served as the control group. The fecal specimens of patients with newly diagnosed tuberculosis were collected from August 2017 to July 2018 in the Xi’an Chest Hospital. Similarly, inclusion was stopped once 30 specimens were collected as the tuberculosis group. The diversity and similarity of gut microbiota in the control group and the tuberculosis group were analyzed by denaturing gradient gel electrophoresis, and the proportion of the bacteria groups in the main bands after polymerase chain reaction-denaturing gradient gel electrophoresis was analyzed.Results The number of bands (13.57±3.37) and the diversity index of the bacteria (H' index; 2.55±0.27) in the control group were significantly higher than those in the tuberculosis group ((8.60±4.19) and 1.99±0.52)), respectively, with the statistically significant differences (t=12.55, P<0.001; t=6.75, P<0.001). A statistically significant difference (Z=-73.38, P<0.001) was found in the intra-group Dice similarity coefficient (Cs) between the control group (22.90% (16.20%, 29.30%)) and the tuberculosis group ((35.35% (23.73%, 44.98%)) (both indicate the median and quartile). Sequencing alignment showed that the main gut microbiota of the control group and the tuberculosis group were mainly Bacteroides, accounting for 73.96% (179/242) and 79.40% (158/199), respectively. The proportion of Prevotella (9.04%, 18/199) in the tuberculosis group was lower than that in the control group (24.38%, 59/242), and the difference was statistically significant (χ ²=17.82, P<0.001). The proportion of Uncultured bacterium genus in the tuberculosis group (11.56%, 23/199) was higher than that in the control group (0, 0/242), and the difference was statistically significant (χ ²=29.51, P<0.001). Conclusion The gut microbiota of healthy people and tuberculosis patients are different due to different immune functions. The occurrence of tuberculosis may result in the changes in the composition ratio of Prevotella and Uncultured bacterium.

Key words: Tuberculosis, Intestines, Bacteria, Polydeoxyribonucleotides, Denaturing gradient gel electrophoresis

杨翰,李欢,郗隽,李爱芳,徐纪茹. 初治结核病患者肠道菌群变性梯度凝胶电泳指纹图谱分析[J]. 中国防痨杂志, 2019, 41(9): 985-992. doi: 10.3969/j.issn.1000-6621.2019.09.013

Han YANG,Huan LI,Jun XI,Ai-fang LI,Ji-ru XU. Fingerprint analysis of gut microbiota by denaturing gradient gel electrophoresis in newly diagnosed tuberculosis patients[J]. Chinese Journal of Antituberculosis, 2019, 41(9): 985-992. doi: 10.3969/j.issn.1000-6621.2019.09.013

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肠道菌群(属/种)	对照组(242株)		结核组(199株)		χ²值	P值
肠道菌群(属/种)	菌株数	构成比(%)	菌株数	构成比(%)	χ²值	P值
Bacteroides 属	179	73.96	158	79.40	1.79	0.215
Bacteroides chinchillae	0	0.00	7	3.52	8.65	0.004
Bacteroides coprocola	12	4.96	7	3.52	0.55	0.491
Bacteroides coprophilus	8	3.31	0	0.00	6.70	0.009
Bacteroides dorei	0	0.00	27	13.57	34.98	<0.001
Bacteroides faecichinchillae	0	0.00	7	3.52	8.65	0.004
Bacteroides faecis	0	0.00	22	11.06	28.16	<0.001
Bacteroides graminisolvens	21	8.68	0	0.00	18.13	<0.001
Bacteroides helcogenes	2	0.83	0	0.00	1.65	0.504
Bacteroides koreensis	0	0.00	6	3.02	7.40	0.080
Bacteroides kribbi	0	0.00	18	9.05	22.82	<0.001
Bacteroides mediterraneensis	2	0.83	0	0.00	1.65	0.504
Bacteroides plebeius	6	2.48	0	0.00	5.00	0.035
Bacteroides sp	0	0.00	5	2.51	6.15	0.018
Bacteroides stercoris	0	0.00	10	5.03	12.44	<0.001
Bacteroides thetaiotaomicron	14	5.79	0	0.00	11.89	<0.001
Bacteroides uniformis JCM5828	51	21.07	0	0.00	47.42	<0.001
Bacteroides vulgatus	63	26.03	42	21.11	1.46	0.261
Bacteroides xylanisolvens	0	0.00	7	3.52	8.65	0.004
Prevotella属	59	24.38	18	9.04	17.82	<0.001
Romboutsia lituseburensis 属	2	0.83	0	0.00	1.65	0.504
Saccharicrinis marinus 属	2	0.83	0	0.00	1.65	0.504
Uncultured bacterium 属	0	0.00	23	11.56	29.51	<0.001
Uncultured bacterium 16sms157-2b03	0	0.00	2	1.01	2.44	0.203
Uncultured bacterium 218004-1-114	0	0.00	3	1.51	3.67	0.091
Uncultured bacterium denovo9345_591_35209	0	0.00	2	1.01	2.44	0.203
Uncultured bacterium G11_1_2c12_	0	0.00	4	2.01	4.91	0.041
Uncultured bacterium OTU561	0	0.00	2	1.01	2.44	0.203
Uncultured bacterium OTU939 16S	0	0.00	3	1.51	3.67	0.091
Uncultured bacterium YO00054E08	0	0.00	5	2.51	6.15	0.018
Uncultured Bacteroidetes bacterium	0	0.00	2	1.01	2.44	0.203