| There are a lot of microbial resources in human stomach environment,which are closely related to human health and diseases.Gastric cancer and gastric ulcer are common digestive tract diseases.So it is very important to understand the diversity of gastric flora and the interaction among the members of microorganisms.Long term infection of Helicobacter pylori is one of the main causes of gastric diseases.To explore the interaction between Helicobacter pylori and gastric flora can provide necessary data support for the treatment and eradication of Helicobacter pylori infection.In this study,gastric mucosa of patients with gastric cancer and gastric ulcer were taken as the main research objects.Based on 16S rRNA gene high-throughput sequencing technology and microbial culture,the structure and diversity of gastric microbial community were analyzed.Secondly,the isolated culturable strains were co cultured with Helicobacter pylori SBK to screen antagonistic strains.Genomic analysis of antagonistic strain Actinomyces timonensis R4-12-8 was carried out,and the inhibition mechanism of the antagonistic strain on Helicobacter pylori SBK was explored through proteomic differences.The results are as follows:1.Isolation and culture of the microorganisms from gastric mucosa samples from patients with gastric cancer and gastric ulcer,and analyse the gastric microbial diversity based on the culture and 16S rRNA gene high-throughput sequencing.A total of 536 strains of bacteria were isolated by pure culture,which includes 10 potential new species.386 strains of bacteria were isolated from gastric cancer samples and distributed in 49 genera and 122 species.150 bacteria were isolated from gastric ulcer samples,which were distributed in 17 genera and 49 species.Consistent with the results of 16S rRNA gene high-throughput sequencing,gastric microorganisms mainly distributed in four phyla:Proteobacteria,Firmicutes,Actinomycetes and Bacteroides.The results of 16S rRNA gene high-throughput sequencing showed that different clinical states shaped different gastric microorganism structures and formed specific bacterial groups.Helicobacter pylori is the dominant group in ulcer tissue,but the abundance of Helicobacter pylori is significantly decreased in gastric cancer samples.In gastric cancer samples,the microbiol groups in tumor tissues and peritumor tissues were significantly different,and the abundance of Helicobacter pylori in tumor tissues decreased significantly.It is suggested that the composition of gastric microorganisms may not be related to the development stage of gastric cancer,but determined by the specific microhabitat of gastric tissue.In gastric ulcer samples,Helicobacter pylori was the dominant.In addition,the difference of microbial community in different human stomach is even more significant.2.Based on the gastric microorganisms obtained by pure culture method,we screened the antagonistic strains of Helicobacter pylori SBKA total of 6 antagonistic strains were screened out from the isolated strains,which distributed in Actinomyces,Streptococcus,Lactobacillus,Aloscardovia,Enterococcus.Actinomyces timonensis R4-12-8 was co cultured with Helicobacter pylori SBK in Transwell chamber.The results showed that Actinomyces timonensis R4-12-8 could induce coccoid conversion of H,pylori cells.We have found that Actinomyces timonensis R4-12-8 produced and released one or more diffusible factors that inhibit the growth of Helicobacter pylori SBK.3.We explored the mechanism of action of antagonistic strains against Helicobacter pylori by genome and proteome analysisThree secondary metabolic pathways were found in the genome of Actinomyces timonensis R4-12-8.Some metabolic pathways were annotated by KEGG database,including metabolism of terpenoids and polyketides,biosynthesis of other secondary metabolites.Helicobacter pylori SBK was co cultured with the supernatant of Actinomyces timonensis R4-12-8,and the strains were collected for proteomic sequencing.A total of 2521 quantifiable proteins were identified.The differentially expressed proteins were mainly distributed in cytoplasm,periplasm,extracellular and outer membrane.The Go database was used for functional enrichment analysis in three aspects,including biological processes,cellular components and molecular functions.To predict the mechanism of action of strain Actinomyces timonensis R4-12-8 on Helicobacter pylori SBK by differentially expressed proteins.Actinomyces timonensis R4-12-8 may promote KDO biosynthesis,inhibit glycolysis and promote chemotaxis of Helicobacter pylori SBK. |
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