Comparative Analysis Of Intestinal Bacterial Diversity In Healthy People,Patients With Non-metastatic Colorectal Cancer And Patients With Colorectal Cancer

Objective:In recent years,a large number of studies have reported that intestinal microbiota affects the occurrence and development of colorectal cancer.In colorectal cancer,the imbalance of intestinal microbiota is the key to the development and progression of colorectal cancer.Microorganisms can change the microenvironment of host cells by regulating metabolites,inducing inflammation,and affecting immune responses,and then promote the transformation of colorectal epithelial cells and the proliferation of tumor cells,therefore accelerate the progress of tumors.The treatment of colorectal cancer includes surgery,radiotherapy and chemotherapy,targeted therapy and immunotherapy.However,nearly half of the patients will eventually have distant metastases such as liver and lung.A profound analysis of the relationship between the intestinal flora and colorectal cancer and its liver metastasis will surely provide new opportunities for studying the mechanism of colorectal cancer and developing treatment methods for the microbiome.16SrDNA sequencing analysis is the most commonly used method in intestinal microbiome research and is used to help determine the correlation between microbial communities and colorectal cancer.This non-invasive method can provide important information on changes in the intestinal ecosystem associated with colorectal cancer.The current research is mainly to compare the difference between the fecal microbiome of healthy people and colorectal cancer patients.Previous studies have compared the fecal microbiome of colorectal cancer patients with the healthy group,and the results show that although there are generally similar microbial communities,a large number of enriched and missing microorganisms can be identified to distinguish between colorectal cancer patients and control groups.This study aims to research the intestinal bacterial diversity of native healthy people,patients with non-metastatic colorectal cancer and patients with colorectal cancer liver metastases,understand the predominant bacteria in the intestine among the three groups of people,and then analyze the intestinal flora structure of patients with non-metastatic colorectal cancer and patients with colorectal cancer liver metastases from the perspective of microorganisms to explore the possible differences in intestinal flora between patients with non-metastatic colorectal cancer and colorectal liver metastasis.This research expects to provide new ideas for the mechanism,diagnosis and treatment of colorectal cancer and colorectal cancer liver metastasisMethods:1.After obtaining the DNA of fecal flora in 8 normal persons,6 patients with non-metastatic colorectal cancer,and 6 patients with colorectal liver metastasis,designing the universal primer 338F-806R by conservative region for PCR amplification and purification to construct the library;2.The PEreads obtained by Miseq sequencing are first spliced according to the overlap relationship,and at the same time,the quality of the sequence is controlled and filtered.After the samples are differentiated,OTU(Operational taxonomic units)cluster analysis and species taxonomy analysis are performed to obtain the corresponding species information and distribution of abundance.3.Based on the OTU clustering analysis results,the OTU conducts a variety of diversity index analysis,as well as the depth of sequencing testing.On the one hand,performing an index group difference test and dilution curve analysis on the Alpha diversity index to obtain species richness and uniformity in the sample degree information.On the other hand,analyzing the sample composition obtained species Venn map,community Bar map and Pie map and community Heatmap map,to obtain the specific differences in microbial composition between different samples,and using Kruskal-Wallis H test and Wilcoxon rank-sum test to conduct a significant analysis test on the difference.Results:1.By comparing the Chaol,ACE,simpson,and Shannon indexes of the normal group,stage patients with non-metastatic colorectal cancer group,and colorectal cancer liver metastasis group,we found that compared with normal people,the number of intestinal microbial species in patients with colorectal cancer liver metastasis is more abundant in terms of species abundance.2.The predominant flora and its abundance of intestinal flora in term of door、class、order、family and genus are different in normal people and colorectal cancer patients,mainly as follow:the level of door:Bacteroidetes、Proteobacteria、Verrucomicrobia;the level of class:Bacteroidale、Gammaproteobacterla、Verrucomicrobiae、Bacilli、Actinobacteria;the level of order:Bacteroidales、Enterobacteriales、Verrucomicroblales;the level of family:Bacteroidaceae、Ruminococcaceae、Enterobacteriaceae、Prevotellaceae、Rikenellaceae、Acidaminococcaceae;the level of genus:Bacteroides、Faecalibacterium、、Escherichia-Shigella、Blautia、Prevotella、Odoribacter、Porphyromonas、Parvimonas、Lachnospira、Parabacteroides、Agathobacter、Alistipes.3.There are certain differences in the abundance of the predominant flora at the levels of phylum,phylum,order,family,and genus between patients with non-metastatic colorectal cancer and patients with colorectal cancer liver metastasis.The predominant flora of patients with non-metastatic colorectal cancer are Bacteroides、Escherichia_Shigella、Blautia、Klebsiella、Agathobacter;The predominant flora of patients with colorectal cancer liver metastasis are Bacteroides、Alistipes、Escherichia_Shigella、Ruminococcus]__gnavus_group.Conclusion:1.Compared with healthy people,the number of intestinal microbial species in patients with colorectal cancer liver metastasis is more abundant.2.The genus Blautia is significantly higher in patients with non-metastatic colorectal cancer than in patients with colorectal cancer liver metastasis,the genus Alistipes is significantly higher in patients with colorectal cancer liver metastasis than patients with non-metastatic colorectal cancer.