PCR-DGGE对新生儿坏死性小肠结肠炎肠道菌落结构及其动态性研究  

Intestinal microbial community diversity in neonatal necrotizing enterocolitis analyzed by PCR-DGGE

作　　者：郑树芳[1] 徐艳珍[1] 艾青[1] 余加林[1] 

机构地区：[1]重庆医科大学附属儿童医院新生儿科/儿童发育疾病研究省部共建教育部重点实验室/儿科学重庆市重点实验室/重庆市儿童发育重大疾病诊治与预防国际科技合作基地,400014

出　　处：《重庆医学》2012年第29期3026-3029,共4页Chongqing medicine

摘　　要：目的探讨新生儿坏死性小肠结肠炎(NEC)患儿肠道菌落的结构和动态性变化。方法从NEC疾病组(n=16例)及对照组(n=16例)的新生儿连续不同时间段的粪便提取肠道细菌总DNA,用357F-GC/518R引物扩增16SrDNA的V3可变区,结合变性梯度凝胶电泳(DGGE)技术分析。结果 NEC组与对照组比较,肠道细菌菌落的Shannon指数相对较高(P<0.05),分布不均,波动明显,细菌的多样性高于对照组,细菌菌落多样性特征存在一定的差异。NEC组的Shannon指数在发病初期明显增多,随着疾病的治疗与恢复,呈先降后升的趋势(Shannon指数从大于或等于50降至小于或等于30以下然后波动回升至35左右),DGGE胶回收测序显示新生儿肠道优势菌群以肺炎克雷伯杆菌属、大肠埃希菌、肠球菌为主,NEC组与对照组细菌种类没有明显的区别,存在数量上的差异,显示了DGGE技术的优越性。结论 NEC患儿的肠道菌群失衡可能是NEC发病细菌感染的重要因素。Objective To analysis intestinal bacterial community composition and dynamics of change in neonatal necrotizing enterocolitis（NEC）. Methods Total DNA of fecal intestinal bacteria was directly extracted from NEC diseases（n= 16） and the control group（n=16） of newborns continuously at different times using the 357F-GC and 518R primers amplified V3 variable region of the 16S rRNA gene,combined with denaturing gradient gel electrophoresis（DGGE）. Results The NEC group compared with the control group, Shannon index of intestinal bacterial was relatively higher（P〈0.05）, the uneven distribution of apparent fluctuations in the diversity of bacteria was higher than the control, there were some differences of bacterial colony diversity characteristics. Shannon index of NEC group increased significantly in the early stage, was down then up trend with the treatment of the disease and recovery（The Shannon index from ≥ 50 to ≤ 30 down and slowly rose to about 35）,and neonatal intestinal advantage flora genus recycling from DGGE were Klebsiella pneumoniae, Escherichia coli, Enterococcus. The main bacterial types of NEC and control groups were no significant difference, there was quantitative differences, showing the superiority of the DGGE technique. Conclusion NEC group existed bacterial imbalance in gut, which might be an important factor in NEC intestinal bacterial infections than a single pathogen.

关 键 词：小肠结肠炎 坏死性 DGGE 肠杆菌科 分子多态性 婴儿 新生 

分 类 号：R722.1[医药卫生—儿科]