Bibiloni R, Simon M A, Albright C, Sartor B, Tannock G W
Department of Microbiology and Immunology, University of Otago, Dunedin, New Zealand.
Lett Appl Microbiol. 2005;41(1):45-51. doi: 10.1111/j.1472-765X.2005.01720.x.
To test combined polymerase chain reaction amplification of 16S rRNA gene sequences and denaturing gradient gel electrophoresis (PCR/DGGE) as an analytical method to investigate the composition of the large bowel microbiota of mice during the development of colitis.
The colonic microbiota of formerly germfree interleukin 10 (IL-10)-deficient mice that had been exposed to the faecal microbiota of specific pathogen-free animals was screened using PCR/DGGE. The composition of the large bowel microbiota of IL-10-deficient mice changed as colitis progressed. DNA fragments originating from four bacterial populations ('Bacteroides sp.', Bifidobacterium animalis, Clostridium cocleatum, enterococci) were more apparent in PCR/DGGE profiles of colitic mice relative to non-colitic animals, whereas two populations were less apparent (Eubacterium ventriosum, Acidophilus group lactobacilli). Specific DNA:RNA dot blot analysis showed that bifidobacterial ribosomal RNA (rRNA) abundance increased as colitis developed.
PCR/DGGE was shown to be an effective method to demonstrate changes in the composition of the large bowel microbiota of mice in relation to progression of inflammatory disease. The intensity of staining of DNA fragments in DGGE profiles reflected increased abundance of bifidobacterial rRNA in the microbiota of colitic animals. As bifidobacterial fragments in PCR/DGGE profiles generated from microbiota DNA showed increased intensity of fragment staining, an increase in bifidobacterial numbers in colitic mice was indicated.
PCR/DGGE analysis demonstrated an altered composition of the large bowel microbiota of colitic mice. This work will allow specific groups of bacteria to be targeted in future research concerning the pathogenesis of colitis.
测试联合聚合酶链反应扩增16S rRNA基因序列和变性梯度凝胶电泳（PCR/DGGE）作为一种分析方法，以研究小鼠结肠炎发展过程中大肠微生物群的组成。
使用PCR/DGGE筛选曾无菌的白细胞介素10（IL-10）缺陷小鼠的结肠微生物群，这些小鼠已接触过无特定病原体动物的粪便微生物群。随着结肠炎的进展，IL-10缺陷小鼠大肠微生物群的组成发生了变化。相对于非结肠炎动物，来自四个细菌群体（“拟杆菌属”、动物双歧杆菌、梭状梭菌、肠球菌）的DNA片段在结肠炎小鼠的PCR/DGGE图谱中更明显，而两个群体则不那么明显（膨胀真杆菌、嗜酸乳杆菌群）。特异性DNA:RNA斑点印迹分析表明，随着结肠炎的发展，双歧杆菌核糖体RNA（rRNA）丰度增加。
PCR/DGGE被证明是一种有效的方法，可证明小鼠大肠微生物群组成与炎症性疾病进展相关的变化。DGGE图谱中DNA片段的染色强度反映了结肠炎动物微生物群中双歧杆菌rRNA丰度的增加。由于从微生物群DNA生成的PCR/DGGE图谱中双歧杆菌片段的染色强度增加，表明结肠炎小鼠中双歧杆菌数量增加。
PCR/DGGE分析表明结肠炎小鼠大肠微生物群的组成发生了改变。这项工作将使特定细菌群体成为未来结肠炎发病机制研究的目标。
