Analysis Of The Influence Of Operation On Intestinal Flora Of Patients With Esophageal Squamous Cell Carcinoma Based On High-throughput Sequencing Technique

Background:Esophageal cancer has become an important problem threatening human life and health because of its hidden early symptoms and strong invasiveness.China is a high incidence area of esophageal cancer,of which more than 90% are esophageal squamous cell carcinoma.At present,the etiology of esophageal cancer is still unclear.In recent years,it has been found that intestinal flora plays an important role in the occurrence and development of esophageal cancer.Surgical resection is still the main treatment for early esophageal cancer,but there is no report on the effect of surgical resection on the changes of intestinal flora.Objective:High-throughput sequencing technique was used to analyze the diversity and structural differences of intestinal flora in fecal samples of patients with esophageal squamous cell carcinoma before and after operation.To explore the influence of operation on intestinal flora of patients with esophageal squamous cell carcinoma by screening differential flora,and to find intestinal microbial markers with diagnostic value for patients undergoing operation,so as to provide theoretical basis for exploring and developing new microbial therapy.Methods:1.18 patients with esophageal squamous cell carcinoma admitted to Nanyang Central Hospital from November,2021 to May,2022 were selected.The stool samples of each patient before and after operation were collected,and the basic information and related clinical data of each patient were recorded.2.In order to explore the composition and structure of intestinal flora in patients with esophageal squamous cell carcinoma before and after operation,a series of microecological data were analyzed,including species composition analysis,species α diversity analysis and β diversity analysis.The specific process is as follows:(1)extracting bacterial DNA;from stool samples;(2)PCR amplification of bacterial genomic DNA,construction of library and 16S rRNA high-throughput sequencing;(3)sequence processing;(4)Species composition analysis and species diversity analysis;(5)Statistical analysis.Results:1.High-throughput sequencing results show that: In this study,36 fecal samples were subjected to 16 s rRNA high-throughput sequencing,and 3,556,410 effective sequences were obtained.After denoising,2,368,085 high-quality sequences were obtained,with an average length of 416.7bp,and most of them were between 404 bp and 430 bp.A total of 29 phyla,70 classes,126 orders,228 families,497 genera and 698 species were obtained.2.The results of species composition analysis show that: The dominant intestinal flora in patients with esophageal squamous cell carcinoma before and after operation are: Firmicutes,Bacteroides,Proteus and Actinomycetes,accounting for more than 99% of the total community abundance;After operation,compared with before operation,the abundance of Actinomycetes increased,while the abundance of Firmicutes,Bacteroides and Proteobacteria decreased.The main dominant flora of intestinal flora in patients with esophageal squamous cell carcinoma before and after operation are: Streptococcus,Shigella,Bifidobacterium,Bacteroides,Lactobacillus,Fecal,Blaut and Roche;After operation,compared with before operation,the abundance of Streptococcus,Bifidobacterium,faecium,Lactobacillus and Blaut increased,while the abundance of Shigella,Bacteroides and Roche decreased.3.The results of species diversity analysis show that:In the analysis of α diversity index,Chao1 index,observed categories index,Shannon index,Simpson index,Faith’s PD index,Pielou’s evenness index and Good’s coverage index showed no significant difference(all P > 0.05),which indicated that the richness,diversity,uniformity and coverage of intestinal flora in patients with esophageal squamous cell carcinoma were similar before and after operation.In β diversity analysis,the test statistic R<0 indicates that the intra-group difference is greater than the inter-group difference,and R>0 indicates that the inter-group difference is greater than the intra-group difference.The results showed that there were differences in intestinal flora between patients with esophageal squamous cell carcinoma before and after operation(P=0.043,R=0.065).4.The results of species differences and marker species analysis show that: In the Venn diagram,2484 ASVs are common before and after surgery,8997 ASVs are unique to patients before surgery and 8574 ASVs are unique to patients after surgery.According to LEfSe analysis,the intestinal microorganisms significantly enriched in patients with esophageal squamous cell carcinoma after operation are: Bifidobacterium,Bifidobacterium,Bifidobacterium,Verona,Microbiology,Fusobacterium,Fusobacterium,Fusobacterium,Fusobacterium and Crescent;The intestinal microorganisms significantly enriched before operation are mainly: bacteria,Bacillus,Vironidae,Massilia,Macromonas and Micromonas.Conclusion:In this study,16S rRNA high-throughput sequencing technique was used to analyze the fecal flora of patients with esophageal squamous cell carcinoma before and after operation.It was found that the diversity and richness of intestinal flora in patients with esophageal squamous cell carcinoma did not change significantly before and after operation,but there were differences in the composition of main intestinal flora before and after operation.After operation,the intestinal flora of Bifidobacterium and Fusobacterium were significantly enriched compared with that before operation.The changes in the composition and structure of intestinal flora may be related to the surgical removal of tumors to prevent the progress of esophageal squamous cell carcinoma.