Establishment Of Animal Model For Infection Of Intestinal Multiple Drug-resistant Bacteria And Study Of Probiotics Intervention

AIM:Exploring the methods of establishment of animal model for infection of intestinal multiple drug-resistant bacteria, to study the protective effect of probiotics in the intestinal infection with multiple drug resistance pseudomonas aeruginosa to intestinal epithelial cells of mice; At the same time, this research judges the intervention function of probiotics in the pathogenic bacteria itself resistance, for the control and treatment of resistant bacteria infection.METHODS:1. Male BALB/C mice were divided into four groups: Normal control group, MDR-PA group, MDR-PA+antibiotics group, MDR-PA+fasting group.2. The infection models of mice were established with antibiotic treatment and by gavage multiple drug-resistant bacteria fluid. Male BALB/C mice were divided into four groups:Normal control group, MDR-PA infection group, MDR-PA+LP group, LP group.3. The changes of mice of method1and method2mental state, feces and weight were observed and recorded during the experimental process at the same time every day. The contents of ileocecal junction were collected for bacterial culture, identification and drug susceptibility experiment. The colonic tissues were collected for pathology observation and inflammation score after the experiment. The expressions of TNF-a and INF-y in the colon were measured at the end of the experiment. The colonic tissues of the method2were collected for pathology observation, inflammation score, permeability, TER and tight junction protein expression detection after the experiment.4. The inhibition experimental research of probiotics in the adherence and invasion of multiple drug resistance pseudomonas aeruginosa:the adhesion experiment included two methods, adhesion inhibition and competitive exclusion experiment. The effect of probiotics on the invasion of pathogens to Caco-2cells was examined by competitive exclusion assays. The apoptosis of the multiple drug resistance pseudomonas aeruginosa inducing Caco-2cells was examined by the adhesion inhibition and competitive exclusion assays, and then the line cell dyeing was examined by flow cytometry. The change of pathogenic bacteria itself drug resistance was examined by bacterial culture and drug sensitive tests.RESULTS:1. Three groups of mice all showed inflammation factor and inflammatory lesions were increased significantly after treatment. The inflammation factors and inflammation score of the MDR-PA+antibiotics group mice increased more obviously than the other two groups. The inflammatory lesions of the MDR-PA+antibiotics group were more serious than the rest of two groups under the microscopic observation.2. The colonic mucosa epithelial cells were observed in all MDR-PA mice in which paracellular permeability was increased, in conjunction with decreased expression and redistribution of ZO-1, occludin, and claudin-1. TNF-a and INF-y concentration had significantly increased. After LP interference, colonic mucosa epithelial cells bypass of the permeability was decreased, tight junction related protein ZO-1, occludin, and claudin-1expressing and distribution were close to normal, and there were statistical significance (P<0.01). Inflammatory factor TNF-α and INF-γ concentration were close to normal, and there were statistical significance (P<0.05). The medicine sensitive experiment results of multiple drug resistance in pseudomonas aeruginosa have changed, but this was not statistically significant.3. The results showed that the probiotics was dose dependent way inhibiting the adhesion and invasion of MDR-PA to Caco-2cells. The aggressivity of MDR-PA decreased by86%when probiotics dose for was come to109CFU. Different doses of MDR-PA and Caco-2cells were incubation2h, using flow cytometry instrument testing, the results found that MDR-PA can induce intestinal epithelial cell apoptosis, along with the increase of the dose of apoptosis rate increased gradually. LP can inhibit the MDR-PA to Caco-2cells induced apoptosis, and the inhibition effect of adhesion inhibition method was more apparent than competitive exclusion method.4. After LP interference, the multiple resistance quality of pseudomonas aeruginosa has changed. Compared with before, the diameter of drug susceptibility in MEM, TZP, ATM-30and CAZ-30were increased, but this was not statistically significant.CONCLUSION:1.Three methods of this experiment all can be successfully to establish animal model with intestinal infection by multiple drug resistance pseudomonas aeruginosa, and each model has its different characteristics.2. Probiotics can treat diarrhea of mice intestinal infection by multiple drug resistance pseudomonas aeruginosa and protect the intestinal barrier function.3. Probiotics can inhibit the adhesion and invasion of MDR-PA to Intestinal epithelial cells.4. Probiotics can weaken the resistance of multiple drug resistance in pseudomonas aeruginosa after LP interference, but this was not statistically significant.