腸道菌群結(jié)構(gòu)變化與結(jié)直腸癌相關(guān)性的研究
發(fā)布時(shí)間：2018-08-17 09:50
【摘要】：目的:通過研究正常人與結(jié)直腸癌患者腸道微生物的多樣性,了解兩類人群腸道優(yōu)勢菌群,推測可能與結(jié)直腸癌的發(fā)生發(fā)展相關(guān)的菌屬;并從微生物的角度分析左、右半結(jié)腸癌的菌群結(jié)構(gòu),探討左右半結(jié)腸癌可能存在的差異。方法:獲取8位正常人,8位右半結(jié)腸癌和5位左半結(jié)腸癌患者糞便菌群DNA后,利用保守區(qū)設(shè)計(jì)通用引物515F-806R進(jìn)行PCR擴(kuò)增并純化,構(gòu)建文庫;使用HiSeq上機(jī)測序后,對原始數(shù)據(jù)進(jìn)行拼接、過濾,獲得有效數(shù)據(jù),基于有效數(shù)據(jù)進(jìn)行OTUs( Operational taxonomic units )聚類和物種分類分析。根據(jù)OTUs聚類結(jié)果,對每個(gè)OTU的代表序列做物種注釋,得到對應(yīng)的物種信息和基于物種的豐度分布情況;同時(shí)對OTUs進(jìn)行豐度、Alpha多樣性計(jì)算、Venn圖和花瓣圖等分析,一方面,獲得了樣品內(nèi)物種豐富度和均勻度信息、不同樣品或分組間的共有和特有OTUs信息等;另一方面,對OTUs進(jìn)行多序列比對并構(gòu)建系統(tǒng)發(fā)生樹,進(jìn)一步比較不同樣品和分組的群落結(jié)構(gòu)差異,并通過PCoA和PCA、NMDS等降維圖和樣品聚類樹進(jìn)行展示。同時(shí),為進(jìn)一步挖掘分組樣品間的群落結(jié)構(gòu)差異,選用T-test以及MetaStat統(tǒng)計(jì)分析方法對分組樣品的物種組成和群落結(jié)果進(jìn)行差異顯著性檢驗(yàn)。結(jié)果:通過對比正常人組與結(jié)直腸癌組的Chao1、ACE、simpson和Shannon指數(shù),發(fā)現(xiàn)與正常人相比,結(jié)直腸癌患者腸道菌群的多樣性相對較低,但是其菌群豐度相對較高。正常人與結(jié)直腸癌患者腸道菌群在門、綱、目、科、屬的優(yōu)勢菌群及其豐度存在差異,主要表現(xiàn)為:在門水平:Firmicutes、Bacteroidetes、Proteobacteria;綱水平:Bacteroidales、Clostridiales、Selenomonadales、Burkholderizles、Enterobacteriales;目水平:Clotridiales、Bacteroidales、Enterobacteriales;科水平 Bacteroidaceae、Lachnospiraceae、Ruminococcaceae、Porphyromonadaceae;屬水平:Bacteroides、Megamonas、Faecalibacterium、Parasutterella、Parabacteroides、Lachnoclostridium、Escherichia-Shigella、Subdoligranulum。結(jié)合系統(tǒng)發(fā)育樹分析,發(fā)現(xiàn)對比正常組,腫瘤組表現(xiàn)為某些菌群豐度增加,如擬桿菌屬、梭桿菌屬、埃希氏菌屬、糞腸球菌、卟啉單胞菌屬等;同時(shí)一些菌群的豐度減少,如乳酸桿菌、雙歧桿菌、羅氏菌屬等。結(jié)論:1.正常人腸道微生物多樣性及群落復(fù)雜程度更高。2.正常人與結(jié)直腸癌患者的腸道菌群在門、綱、目、科、及屬水平的豐度上存在著一定差異。一些菌屬在腫瘤患者中豐度較高,可能在腫瘤的發(fā)展過程中起到了推動(dòng)作用,如擬桿菌屬、梭桿菌屬、埃希氏菌屬等;另一些菌屬在腫瘤患者中豐度較低,他們對正常的機(jī)體可能有益處,如乳酸桿菌、雙歧桿菌、羅氏菌屬等。3.左右半結(jié)腸癌患者的腸道優(yōu)勢菌群菌群在門、綱、目、科、及屬水平的豐度上存在著一定差異。右半結(jié)腸癌患者屬水平的優(yōu)勢菌群為Fusobacteria,Bacteroidetes,Lachnospiraceae,Enterobacteriaceae;左半結(jié)腸癌患者屬水平的優(yōu)勢菌群為 Bacteroidetes,B. Fragilis,Lachnospiraceae,Fusobacteria;腸道菌群在左、右半結(jié)腸癌腸道的生物學(xué)特性可能存在差異。
[Abstract]:Objective: To study the diversity of intestinal microorganisms in normal people and patients with colorectal cancer, to understand the dominant intestinal flora of the two groups of people, and to speculate the bacterial genus which may be related to the occurrence and development of colorectal cancer. The DNA of stool flora from 8 normal subjects, 8 right colon cancer patients and 5 left colon cancer patients was amplified and purified by PCR using a universal primer 515F-806R designed by conservative region, and the library was constructed. Clustering and species classification analysis. According to the OTUs clustering results, the representative sequences of each OTU are annotated to get the corresponding species information and the species abundance distribution based on the species. Meanwhile, the OTUs are analyzed by abundance, Alpha diversity calculation, Venn diagram and petal diagram. On the one hand, the species richness and evenness information in the sample is obtained. On the other hand, OTUs were sequentially aligned and phylogenetic trees were constructed to further compare the differences of community structure between different samples and groups, and then displayed by dimensionality reduction maps such as PCoA, PCA, NMDS and sample clustering tree. Results: By comparing the Chao1, ACE, Simpson and Shannon indices of normal and colorectal cancer patients, it was found that the diversity of intestinal flora in colorectal cancer patients was lower than that in normal subjects, but the diversity of intestinal flora was lower. The dominant intestinal flora and its abundance in the phylum, class, order, family and genus of normal people and patients with colorectal cancer are different, mainly as follows: at the phylum level: Firmicutes, Bacteroidetes, Proteobacteria; class level: Bacteroidales, Clostridiales, Selenomonadales, Burkholderizles, Enterobacteriales; class level: Firmicutes, Bacteroidetes, Bacteroidetes, Proteobacteria; class level: Bacteroidales, Clostridiales, Selenomonadales, Burkholderiz Tridiales, Bacteroidales, Enterobacteriales; Family level Bacteroidaceae, Lachnospiraceae, Ruminococcaceae, Porphyromonadaceae; Genus level: Bacteroides, Megamonas, Faecalibacterium, Parasutterella, Parabacteroides, Lachnoclostridium, Escherichia-Shigella, Doligranulum. In group B, the abundance of some bacteria increased, such as Bacteroides, Fusobacterium, Escherichia coli, Enterococcus faecalis, Porphyromonas, and so on. At the same time, the abundance of some bacteria decreased, such as Lactobacillus, Bifidobacterium, Rochella and so on. Conclusion: 1. The intestinal microbial diversity and community complexity of normal people and colorectal cancer were higher. There are certain differences in the abundance of intestinal flora among cancer patients at the phylum, class, order, family, and genus levels. Some bacteria are abundant in cancer patients and may play a role in promoting the development of cancer, such as Bacteroides, Fusobacterium, Escherichia, etc. Others are low in cancer patients and they are normal to them. There are some differences in the abundance of the dominant flora in the intestine between the right and left hemicolonal cancer patients at the phylum, class, order, family, and genus levels. The dominant flora in the right hemicolonal cancer patients at the genus level are Fusobacteria, Bacteroidetes, Lachnospiraceae, Enterobacteriaceae. Bacteroidetes, B. Fragilis, Lachnospiraceae, Fusobacteria were the predominant flora in the genus of hemicolonal cancer patients, and the biological characteristics of the left and right colon cancer may be different.
【學(xué)位授予單位】：昆明醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R735.34
本文編號：2187232
[Abstract]:Objective: To study the diversity of intestinal microorganisms in normal people and patients with colorectal cancer, to understand the dominant intestinal flora of the two groups of people, and to speculate the bacterial genus which may be related to the occurrence and development of colorectal cancer. The DNA of stool flora from 8 normal subjects, 8 right colon cancer patients and 5 left colon cancer patients was amplified and purified by PCR using a universal primer 515F-806R designed by conservative region, and the library was constructed. Clustering and species classification analysis. According to the OTUs clustering results, the representative sequences of each OTU are annotated to get the corresponding species information and the species abundance distribution based on the species. Meanwhile, the OTUs are analyzed by abundance, Alpha diversity calculation, Venn diagram and petal diagram. On the one hand, the species richness and evenness information in the sample is obtained. On the other hand, OTUs were sequentially aligned and phylogenetic trees were constructed to further compare the differences of community structure between different samples and groups, and then displayed by dimensionality reduction maps such as PCoA, PCA, NMDS and sample clustering tree. Results: By comparing the Chao1, ACE, Simpson and Shannon indices of normal and colorectal cancer patients, it was found that the diversity of intestinal flora in colorectal cancer patients was lower than that in normal subjects, but the diversity of intestinal flora was lower. The dominant intestinal flora and its abundance in the phylum, class, order, family and genus of normal people and patients with colorectal cancer are different, mainly as follows: at the phylum level: Firmicutes, Bacteroidetes, Proteobacteria; class level: Bacteroidales, Clostridiales, Selenomonadales, Burkholderizles, Enterobacteriales; class level: Firmicutes, Bacteroidetes, Bacteroidetes, Proteobacteria; class level: Bacteroidales, Clostridiales, Selenomonadales, Burkholderiz Tridiales, Bacteroidales, Enterobacteriales; Family level Bacteroidaceae, Lachnospiraceae, Ruminococcaceae, Porphyromonadaceae; Genus level: Bacteroides, Megamonas, Faecalibacterium, Parasutterella, Parabacteroides, Lachnoclostridium, Escherichia-Shigella, Doligranulum. In group B, the abundance of some bacteria increased, such as Bacteroides, Fusobacterium, Escherichia coli, Enterococcus faecalis, Porphyromonas, and so on. At the same time, the abundance of some bacteria decreased, such as Lactobacillus, Bifidobacterium, Rochella and so on. Conclusion: 1. The intestinal microbial diversity and community complexity of normal people and colorectal cancer were higher. There are certain differences in the abundance of intestinal flora among cancer patients at the phylum, class, order, family, and genus levels. Some bacteria are abundant in cancer patients and may play a role in promoting the development of cancer, such as Bacteroides, Fusobacterium, Escherichia, etc. Others are low in cancer patients and they are normal to them. There are some differences in the abundance of the dominant flora in the intestine between the right and left hemicolonal cancer patients at the phylum, class, order, family, and genus levels. The dominant flora in the right hemicolonal cancer patients at the genus level are Fusobacteria, Bacteroidetes, Lachnospiraceae, Enterobacteriaceae. Bacteroidetes, B. Fragilis, Lachnospiraceae, Fusobacteria were the predominant flora in the genus of hemicolonal cancer patients, and the biological characteristics of the left and right colon cancer may be different.
【學(xué)位授予單位】：昆明醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：R735.34
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