溃疡性结肠炎患者粪便菌群和乙状结肠黏膜菌群分析

Analysis of bacterial flora in faeces and mucosa of sigmoid colon in patients with ulcerative colitis

目的探讨UC患者与健康对照者粪便菌群和乙状结肠黏膜菌群的差异，以及肠道菌群改变在UC发病中的可能作用及意义。方法以梭菌属、肠球菌属、乳杆菌属、双歧杆菌属及拟杆菌属、埃希菌属作为目标菌群，采用实时荧光定量PCR方法检测35例UC患者（活动期组20例、缓解期组15例）及20名健康体检者（健康对照组）的粪便标本及乙状结肠黏膜活组织检查标本中6种目标菌群的含量。采用两组独立样本t检验比较粪便菌群和乙状结肠黏膜相关菌群的差异；采用方差分析比较对照组、活动期组及缓解期组粪便和乙状结肠黏膜菌群的差异。结果健康对照组粪便菌群中梭菌属、拟杆菌属、肠球菌属、乳杆菌属含量分别为（10．94±0．29）、（12．42±0．39）、（8．73±0．84）、（9．05土0．35）lg拷贝／g，均高于乙状结肠黏膜[分别为（9．36±0．66）、（9．88±0．82）、（7．70±1．17）、（7．96±0．68）lg拷贝／g，t=9．83、12．51、3．20、6．35，P均〈o．05]；埃希菌属含量低于乙状结肠黏膜[（8．03±1．02）lg拷贝／g比（8．91±0．52）lg拷贝／g，t=-3．44，P〈0．05]；粪便菌群与乙状结肠黏膜菌群中双歧杆菌属含量差异无统计学意义[（9．54±0．79）lg拷贝／g比（9．42±0．98）lg拷贝／g，P〉o．05]。UC患者粪便菌群中梭菌属、拟杆菌属、肠球菌属、乳杆菌属、双歧杆菌属含量分别为（9．62±1．13）、（11．31±0．71）、（9．33±0．65）、（8．42±0．80）、（8．85±0．73）Ig拷贝／g，均高于乙状结肠黏膜[分别为（9．00±0．79）、（8．80士0．66）、（7．46±0．82）、（6．82±1．07）、（8．40±0．72），t=2．66、15．28、10．58、7．12、2．56，P均〈0．05]；埃希菌属含量低于乙状结肠黏膜[（8．50±0．52）lg拷贝／g比（9．26±0．87）lg拷贝／g，t=-4．45，P〈0．05]。与健康对照组比较，活动期组粪便菌群中梭菌属、乳杆菌属、双歧杆菌属、拟杆菌属含量[分别为（8．83±0．81）、（7．48±1．59）、（8．55±0．79）、（11．11±0．88）lg拷贝／g]均降低（F=84．45、14．58、10．43、24．91，P均〈O．05），肠球菌属、埃希菌属含量[（9．63±0．39）、（8．74±0．53）lg拷贝／g升高（F=9．87、5．55，P均〈O．05]；而缓解期组仅表现为拟杆菌属含量下降[（11．56±0．21）Ig拷贝／g，P〈O．05]。与健康对照组比较，活动期组乙状结肠黏膜菌群中梭菌属、拟杆菌属、乳杆菌属、双歧杆菌属含量[分别为（8．524-0．30）、（8．34±0．29）、（6．29±0．52）、（8．06±0．21）lg拷贝／g均减少（F=16．99、35．98、22．28、16．08，P均〈O．05]，埃希菌属含量[（9．68±0.56）lg拷贝／g]增加（F=11．19，P〈O．05），肠球菌属含量[（7．19±0．32）lg拷贝／g]差异无统计学意义（P〉0．05）；缓解期组拟杆菌属和双歧杆菌属含量[（9．42±o．48）lg拷贝／g和（8．87±o．89）lg拷贝／g，P均〈0．05]减少，其他菌群含量差异均无统计学意义（P均〉0．05）。无论是粪便菌群还是黏膜相关菌群，活动期组双歧杆菌与肠杆菌比值（B／E值）均〈1（O．98±0．13、0．84±0．05），较健康对照组（1．21±0．19、1．06±0．08）明显下降（F=12．64、76．20，P均〈0．05）。缓解期组B／E值均〉1（1．14±0．08、1．02±0．04），与健康对照组差异无统计学意义（P〉0．05）。结论健康体检者与UC患者粪便和乙状结肠中各种目标菌群的分布均不相同。UC患者粪便菌群和黏膜相关菌群较健康体检者存在明显变化，以活动期组更显著。

Objective To explore the difference in bacterial flora loetween faeces and mueosa of sigmoid colon, the possible role and significance of microbiota alteration in the genesis of ulcerative colitis （UC）. Methods Fusobacterium, Enterococcus, Lactobacillus, Bi fidobacteriurn, Bacteraides and Escherichia were selected as target bacteria colonies. The content of six target bacteria colonies in faeces and mucosa of sigmoid colon of 35 UC patients （20 active UC, and 15 UC in remission） and 20 health controls were detected by real-time fluorescent quantitative polymerase chain reaction （RT-PCR）. Two independent samples t-test was performed to compare the differences in bacterial flora between faeces and mucosa of sigmoid colon. Variance analysis was used to compare the differences in bacterial flora among health controls group,active stage group and remission stage group. Results In health control group, the contents of Fusobacteriurn, 13acteroides, Enterococcus and Lactobacillus in faeces （（10. 94 ± 0. 29）, （12.42±0.39）, （8. 73±0.84）, （9.05 ± 0. 35）, respectively） were higher than those in the mucosa of sigmoid colon （（9.36±0.66）, （9.88±0.82）, （7.70±1.17） and （7.96±0.68）, respectively, t=9.83, 12.51, 3.20 and 6.35, all P〈0.05）. However, the content of Escherichia was lower in faeces than that in the mucosaofsigmoid colon （（8.03±1.02） lg copy/g vs （8.91±0.52） lg copy/g, t=-3.44, P〈 0.05）. There was no difference in the content of Bifidobacterium between faeces and mucosa of sigmoid colon （（9. 54±0.79） lg copy/g vs （9.42±0.98） lg copy/g, P〈0.05）. For UC patients, the contents of Fusobacteriurn, Bacteroides, Enterococcus, Lactobacillus and Bifidobacterium in faeces （ （9.62 ± 1.13）, （11.31±0.71）, （9.33±0.65）, （8.42±0.80） and （8.85±0.73） lg copy/g, respectively） were higher than those in the mucosa of sigmoid colon （（9.00±0.79）, （8.80±0.66）, （7.46±0.82）, （6.82±1.07） and （8.40±0.72） lg copy/g, respectively, t=2.66, 15.28, ]0.58, 7.12 and 2.56, all P〈0.05）. The content of Escherichia was lower in faeces than that in the mucosa of sigmoid colon （（8. 50 ± 0. 52） lg copy/g vs （9.26±0.87） lg copy/g, t= -4.45, P〈0.05）. Compared with health control group, the content of Fusobacterium, Lactobacillus, Bi fidobacterium and Bacteroides （（8. 83 ± O. 81）, （7. 48± 1.59）, （8.55±0.79） and （11.11±0.88） lg copy/g） all decreased （F=84.45, 14.58, 10.43 and 24.91, all P〈0.05）, while the contents of Enterococcus and Escherichia increased （（9.63±0.39） and （8.74± 0.53） lg copy/g, F= 9. 87 and 5. 55, both P〈0.05）. For remission stage group, only the content of Bacteroides decreased （（11.56±0.21） lg copy/g, P〈0.05）. Compared with health control group, the contents of Fusobacterium, Bacteroides, Lactobacillus, Bifidobacterium （（8. 52 ± 0. 30）, （8. 34 ±0.29）, （6.29±0. 52） and （8. 06±0. 21） lg copy/g） all decreased in active stage group （F=16. 99, 35.98,22.28 and 16.08, all P〈0.05）, the content of Escherichia increased （（9.68±0.56） lg copy/g, F=11. 19,P〈0.05）; there was no difference in the content of Enterococcus （（7. 19±0.32） lg copy/g, P〉0.05）. In remission stage group, the contents of Bacteroides fragilis and Bifidobacterium decreased （（9.42±0.48） lg copy/g and （8.87±0.89） lg copy/g, both P〈0.05）, there was no difference in other bacterias （all P〈0.05）. In both faeces and mucosa of sigmoid colon, the ratios of Bifidobacterium and Enterobacteriaceae （B/E value） in active stage group were less than 1 （0. 98±0. 13 and 0. 84±0. 05）, which significantly decreased compared with health control group （1.21±0.19, 1.06±0.08; F=12.64, 76.20, both P〈0. 05）. In remission stage group, B/E values were higher than 1 both in faeces and mucosa （1.14±0.08 and 1.02±0.04）, and there was no difference compared with those of control group （P〉0.05）. Conclusions The distribution of target bacteria in feces and sigmoid colonis differed between health controls and UC patients. There are obvious changes in fecal and mucosa associated bacterial flora in UC patients especially in active stage compared with healthy controls.