功能性便秘儿童肠道菌群DNA的PCR—SSCP和BOX—PCR指纹图谱分析及临床意义

PCR-SSCP and BOX-PCR finger-print analysis of the intestinal flora in patients with childhood functional constipation

目的探讨功能性便秘（functional constipation．FC）儿童肠道微生态菌落的构成及其在便秘发生过程中的作用。方法研究对象分为便秘组和对照组．便秘组15例，平均年龄6．5岁．对照组35例，平均年龄6岁。取所有儿童粪便提取细菌基因组DNA，应用基于细菌16SrDNA的PCR—SK2P技术和基于细菌基因组的BOX—PCR技术对肠道菌落的DNA指纹图谱分析，结合克隆测序和生物信息学技术对FC儿童的肠道菌落进行初步鉴定，并对其组成结构进行分析。结果对照组和便秘组的所有样品分别进行3次以上的PCR—SSCP和BOX—PCR指纹图谱分析，得到了稳定可重复的罔谱。便秘组菌种的种类较对照组明显增多．差异有统计学意义；对照组优势菌主要为拟杆菌属、埃希氏菌属、双歧杆菌属、乳杆菌属；便秘组的优势菌主要为脆弱拟杆菌属、埃希氏菌属、真杆菌属和梭菌属。结论功能性便秘儿童肠道菌群的菌属多样性增加．各肠道菌属间存在比例失调．肠道微生态失调可能参与了便秘的产生或加重过程。

Objective To investigate the bacterial species of the intestinal flora in patients with childhood functional constipation （FC）, and to explore its pathophysiological role in childhood FC. Methods Fifty patients were recruited into this study, including 15 FC patients and 35 controls. Bacterial genome DNA was extracted from stool samples, the DNA finger print was analyzed by PCR-SS- CP and BOX PCR. The bacterial species were identified by DNA finger prints combined with clone-sequence and GenBank database analysis. Results Stable and repeatable intestinal flora finger prints were obtained by PCR SSCP and BOX-PCR. Compared to control group, more bacterial species were found in FC patients. Predominant bacterial species of control group were Bacteroides, Escbericbia, Bifidobacterium and Acidobacterium; Predominant bacterial species of FC group were Bacteroides fragills, Escherichia, Eubacterium and Clostridia. Conclusions The disproportion of predominant bacterial species characterizes the intestinal flora of FC children, which may contribute to initiate or aggra vate FC.