Study On Pathogenicity Mechanism Of Microbial Community Based On The Macro Genomics And Culturomics In Patients With Cholecystitis

ObjectiveThe high-throughput whole genome sequencing and high-throughput separation culture were carried out on the feces of healthy people,patients with cholecystitis and the corresponding bile samples by using the methods of metagenomics and culturomics,and the related bacterial groups of cholecystitis were comprehensively analyzed to explore the pathogenesis of cholecystitis flora.MethodFecal samples were collected from 5 healthy people and 5 cholecystitis patients and corresponding bile samples from these 5 cholecystitis patients.Eleven different media were used for large-scale isolation and culture of the culture group samples and total bacterial DNA was extracted simultaneously.The isolated and cultured bacteria were identified by MALDI-TOF time-of-flight mass spectrometry combined with 16 S r RNA.The samples of the sequencing group were sequenced by the shotgun method and the total DNA in the environment was extracted.Based on the results of culturomics and metagenomics,the pathogenic mechanism of cholecystitis flora was discussed.Results1.727 strains of bacteria,belonging to 22 genera and 40 species,were isolated and identified from 5 fecal samples of healthy people and 5 fecal and bile samples of patients with cholecystitis.Among them,328 strains of bacteria were isolated from the feces of healthy people,belonging to15 genera and 20 species in total.The most commonly isolated strains were Bifidobacterium,Lactobacillus and Escherichia,53,44 and 33 strains respectively.A total of 399 strains of bacteria belonging to 18 genera and 35 species were isolated from patients with cholecystitis.Among them,25 strains of bacteria were isolated from bile samples,belonging to 6 genera and 8 species.The top three strains were Enterococcus,Enterococcus casseliflavus and Klebsiella.374 strains were isolated from fecal samples,belonging to 18 genera and 35 species in total.Among them,the most common genera were Enterococcus,Bacteroides and Escherichia,127,49 and 38 strains were isolated,respectively.The top three strains of fecal samples were Enterococcus,Enterococcus casseliflavus and Escherichia coli.2.High-throughput sequencing was used to detect drug-resistant genes in important strains identified in bile and fecal samples from patients with cholecystitis,we detected 472 genes at different locations in bile samples,and found that 118 genes were resistant to 15 different types of antibiotics,among which arc A gene was the most frequently detected,showing multidrug resistance.A total of 99 genes were detected in different locations of fecal samples.39 genes were found to be resistant to 10 antibiotics,and tet Q gene was the most frequently detected gene,but only to tetracycline.The number of common drug resistance genes detected in the two samples was 18.Conclusion1.There is a strong correlation between the number of Enterococcus in the intestine of patients with cholecystitis and the bile flora.2.There is a potential relationship between gallbladder inflammation and intestinal flora.3.arc A is a multidrug-resistant gene,while tet Q is only resistant to tetracycline.The number of drug-resistant genes in bile and feces samples of patients with cholecystitis was 18,with an overlap rate of15.25 %,which is quite different.4.Based on the results of metagenomics and culturomics,we found that there was a significant difference in the overlapping rate between bile samples and fecal samples,which were 9.09 % and 26.09 %respectively.