新生儿肠道菌群分布特征及其影响因素

发布时间：2018-01-18 09:46

本文关键词：新生儿肠道菌群分布特征及其影响因素　出处：《上海交通大学》2015年硕士论文　论文类型：学位论文

【摘要】：第一部分DGGE分析新生儿期肠道菌群多样性及影响因素目的:利用PCR-DGGE技术,研究新生儿肠道菌群多样性的动态特征,分析影响肠道菌群多样性的相关因素。方法:采集157例足月儿和72例早产儿生后第3、5、7天及3周的粪便标本,提取细菌总DNA,对其16S r DNA的V3可变区进行PCR扩增,扩增产物经DGGE电泳后计算丰富度和Shannon指数,进行统计学分析。结果:(1)新生儿肠道菌群随时间动态变化,3天到3周的丰富度均随时间增多(6.17±2.23,10.34±2.60,13.04±3.82,13.86±3.81),1周内的Shannon指数随时间上升(1.33±0.34,1.74±0.25,1.80±0.31),第3周较第7天的多样性无明显变化(P0.05)。第5天丰富度及Shannon指数明显高于第3天(P0.001)。(2)出生方式的影响:剖宫产儿生后第3天的丰富度和Shannon指数(5.63±2.05,1.24±0.35)明显低于顺产儿(6.82±2.28,1.44±0.30,P0.001);第5天Shannon指数也低于顺产儿(P0.05),1周和3周两组无差异。(3)喂养方式的影响:1周和3周时母乳喂养Shannon指数(1.75±0.32,1.79±0.32)较非母乳喂养低(1.88±0.29,1.92±0.35,P0.05),丰富度无差异(P0.05)。(4)胎龄的影响:足月儿3、5、7天的丰富度和3、5天的Shannon指数均明显高于早产儿。第3周两组无明显差异。结论:新生儿期肠道菌群多样性随时间呈动态性变化,3-5天为菌群发展最迅速时期。剖宫产新生儿早期的肠道菌群多样性低于顺产儿。母乳喂养多样性低于非母乳喂养。早产儿肠道菌群多样性低下,定植延迟。第二部分Illumina高通量测序分析胎龄及喂养方式对早产新生儿期肠道菌群分布特征的影响目的:应用Illumina高通量测序技术,研究胎龄及喂养方式对早产儿肠道菌群分布特征的影响。方法:(1)选取剖宫产、母乳及人工喂养的早产儿35名入喂养组;选取剖宫产、人工喂养早产儿22名入胎龄组。(2)将粪便细菌DNA等浓度等比例混合,扩增16Sr DNA V3区,应用Illumina Miseq平台测序,分析肠道菌群组成特征。结果:(1)早产儿肠道菌群以两个细菌门类--Firmicutes、Proteobacteria、三个纲--Bacilli、Clostridia、Gammaproteobacteria为主。在科水平以Streptococcaceae、Veillonellaceae、Enterobacteriaceae占主要优势。整个观察期未检出Bacteroidetes和Bifidobacterium。(2)喂养方式的影响:两种喂养方式对主要菌门和菌属的相对丰度的影响无明显差异。低丰度水平上,母乳喂养儿比人工喂养儿拥有更丰富的物种,研究发现Lactobacillus母乳组显著高于人工喂养组(P0.05)。而Escherichia甚至在人工喂养组未能检出。另外,在不同时间点两组菌群结构也存在差异,门水平的差异主要表现在3-5天。(3)胎龄的影响:大胎龄组较小胎龄组拥有更丰富的物种,优势菌属Veillonella在大胎龄组明显高于小胎龄组(P0.05)。结论:高通量测序揭示了早产儿肠道菌群特征:更多兼性厌氧菌定植,严格厌氧菌定植延迟。胎龄是影响早产儿肠道菌群分布特征的重要因素。喂养方式对主要菌群结构影响不大,人工喂养对部分低丰度菌群的建立存在不良影响。
[Abstract]:Part I DGGE analysis of intestinal microflora diversity and its influencing factors during neonatal period objective: to study the dynamic characteristics of intestinal microflora diversity in neonates by using PCR-DGGE technique. Methods: the fecal specimens of 157 term infants and 72 preterm infants were collected on the 7th and 3rd week after birth to extract total bacterial DNA. The V3 variable region of 16s r DNA was amplified by PCR. The richness and Shannon index were calculated by DGGE electrophoresis. Results the intestinal microflora of newborns changed dynamically with time from 3 days to 3 weeks. The richness of intestinal microflora was 6.17 卤2.23 卤10.34 卤2.60 with the increase of time. The Shannon index of 13.04 卤3.82v 13.86 卤3.81g per week increased by 1.33 卤0.34 卤1.74 卤0.25 with time. 1.80 卤0.31). There was no significant change in diversity between the 3rd week and the 7th day. The richness and Shannon index on the 5th day were significantly higher than that on the 3rd day (P 0.001. 0. 0. 2). The influence of birth style: the richness and Shannon index were 5.63 卤2.05 on the 3rd day after cesarean section. 1.24 卤0.35) was significantly lower than that in the normal delivery group (6.82 卤2.28) 1.44 卤0.30 (P 0.001); On the 5th day, the Shannon index was also lower than that of the normal birth baby (P0.05). There was no difference between the two groups at 1 and 3 weeks.) the Shannon index of breast-feeding was 1.75 卤0.32 at 1 and 3 weeks after 1 and 3 weeks. 1.79 卤0.32) was lower than that of non-breast feeding (1.88 卤0.291.92 卤0.35 P0.05). The effect of gestational age on the richness was not different (P0.05. 4): the richness and 3. The Shannon index of 5 days was significantly higher than that of preterm infants. There was no significant difference between the two groups at the third week. Conclusion: the diversity of intestinal flora in neonatal period showed dynamic changes with time. The diversity of intestinal flora in the early stage of cesarean section was lower than that in the normal birth, the diversity of breast-feeding was lower than that of non-breast-feeding, and the diversity of intestinal flora of premature infants was low. Second part Illumina high-throughput sequencing was used to analyze the effects of gestational age and feeding mode on the distribution of intestinal flora in preterm newborn. Objective: to apply Illumina high-throughput sequencing technique. To study the effect of gestational age and feeding methods on the distribution of intestinal flora of premature infants. Methods 35 premature infants with cesarean section, breast milk and artificial feeding were selected into the feeding group. The fecal bacterial DNA concentration was mixed to amplify 16Sr DNA V3 region by cesarean section and artificial feeding of 22 premature infants in gestational age group. Illumina Miseq platform was used to analyze the composition of intestinal flora. Results the intestinal flora of preterm infants was divided into two groups: Firmicutes. Proteobacteria, three classes Bacillian Clostridia. The main Gammaproteobacteria is Streptococcaceaeus veillonellaceae. Enterobacteriaceae was dominant. Bacteroidetes and Bifidobacterium. 2 were not detected during the observation period. The effects of two feeding methods on the relative abundance of the main phylum and genus were not significantly different. Breast-feeding babies are richer in species than artificially fed ones. The study found that the Lactobacillus breast milk group was significantly higher than the artificial feeding group P0. 05, but Escherichia was not even detected in the artificial feeding group. In addition. At different time points, there were differences between the two groups, and the difference of gate level was mainly manifested in the influence of gestational age of 3-5 days. The large gestational age group had more abundant species than the small gestational age group. The dominant Veillonella was significantly higher in the large gestational age group than in the small gestational age group (P 0.05). Conclusion: high throughput sequencing revealed the characteristics of intestinal flora of premature infants: more facultative anaerobes were colonized. Strict anaerobe colonization delay. Gestational age is an important factor affecting the distribution of intestinal flora of premature infants. Feeding mode has little effect on the main flora structure, but artificial feeding has a negative effect on the establishment of some low-abundance microflora.
【学位授予单位】：上海交通大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R722.1

【相似文献】