The Study Of Lactobacillus Species In Feces Of Patients With Ulcer Ative Colitis

Chapter One:The analysis of11species of Lactobacilli in the faeces of patients with Ulcerative ColitisObject:In order to analysis the alteration of lactobacillus species in the faeces of patients with ulcerative colitis, we have investigated the composition and quantification of fecal lactobacillus species in use of species-specific primers based on the16S-23S DNA sequence information.Methods:45patients met the diagnosis criteria were selected as observation group, also45healthy people were included as control group. Selective culture were used to detect changes in8groups of bacteria. The lactobacillus species were identified with species-specific primers. Polymerase chain reaction was used to amplify bacterial DNA from faeces and electrophoresis was used to detect the products. Also relative quantitative real-time polymerase chain reaction was use together to determine the distribution and quantification of lactobacillus species of fecal bacteria.Results:In patients with ulcerative colitis, the fecal bacteria were mainly composed with Bifidobacteria, E. coli, Staphylococcus aureus and Enterococcus, also small quantity of Bacteroid and Yeast. Compared with control group, lactobacillus reduced while enterococcus increase. The quantity of Yeast, Bacteroides, Bifidobacterium, E.coli, Staphylococcus aureus and total aerobic bacteria were close to the control group. The detection of11species of lactobacillus showed that the detection rate was lower in observation group in the species of L. crispatus, L. fermentum, L gasseri, and L. salivarius, while another two species increased in detection rate:L. delbrueckii, L. paracasei. There were five species close to normal control in detection rate:L. gallinarum, L. acidophilus, L. casei, L. plantarum, and L. reuteri. The relative quantitative test results showed that fecal lactobacillus including L. crispatus, L. delbrueckii, L. reuteri and E. faecalis were increased according to the control group, while L. gallinarum, L. fermentum, L. johnsonii, L. salivarius, L. acidophilus, L. casei, L. paracasei, L. plantarum and lactobacillus total quantity were reduced in observation group.Conclusion:In patients with ulcerative colitis, the quantity of lactobacillus was reduced while enterococci increased in culturable bacteria in fecal flora. In the species analysis of lactobacillus, the composition was different from normal contral. Relative quantitative analysis detected that3species of lactobacillus and Enterococcus faecalis were increased in patients, while the remaining8species of lactobacillus and lactobacillus total quantity were reduced. Chapter Two:The therapeutic effects of three species of lactobacillus and Enterococcus faecalis on DSS-induced experimental colitis in Balb/c miceObject:According to the results of fecal flora analysis in patients with ulcerative colitis, we selected the appropriate strains of lactobacillus for the treatment of experimental colitis in order to establish the relationship between lactobacillus species and the disease.Methods:A total of80Balb/c mice were enrolled in this experiment, specific pathogen free,6-8-week-old females, free drink of DSS to form the colitis mouse model for the study. The strains of bacteria used in this research were selected according to the results of fecal analysis. Mesalazine was used as positive control agent. We observed the general conditions of mice, survival ratio, fecal properties, fecal occult blood and hemafecia, measurement of colon lengths, DAI scores and pathological scores.Results:The results showed that L. fermentum could reduce the death ratio and improve weight loss than blank control, positive control, L. plantarum group and Enterococcus faecalis group in mice with experimental colitis. It also prevent colon shortening, reduce disease activity, reduce histological score and crypt damage, improve the inflammation damage in experimental colitis. However, L. crispatus seems to reduce the survival ratio and accelerate weight loss in mice, although it didn't aggravate the colonic shortening, it "still increased colonic inflammation damage. L. plantarum and Enterococcus faecalis had no significant treatment in experimental colitis, and Enterococcus faecalis may reduce crypt damage.Conclusion:Through the detection of weight loss, colon length, DAI scores, pathological scores and comprehensive evaluation, we concluded that:L. fermentum showed effective treatment close to mesalazine in DSS-induced experimental colitis in mice. Chapter Three:The study of microbiological characteristics of Lactobacillus fermentumObject:To study the main microbiological characteristics of Lactobacillus fermentum which showed therapeutic effect on experimental colitis.Methods:The study used L. fermentum strain CCTCC M206120, including morphology analysis through light microscopy and electron microscopy, growth characteristics analysis by oxygen tolerance test and resistance to bile acid properties in simulation environment, antibiotic susceptibility analysis through drug sensitivity test, genetic characteristics analysis through the test of G+C ratio and16S DNA sequence analysis.Results:The results showed that the strain of L. fermentum was Gram-positive, facultative anaerobic bacteria, it prefer anaerobic conditions, had the ability of acid and bile tolerance, highly sensitive to32kinds of antibiotics, and its biochemical and genetic characteristics conformed to Lactobacillus fermentum.Conclusion:Preliminary analysised the main microbiological characteristics of L. fermentum strain CCTCC M206120.