The Study Of Severe Risk Factor And Probiotics Intervention Mechanisms In Hospital-acquired Clostridium Difficile Infection

Part 1 Molecular Epidemiology and risk factor for severity of hospital-acquired Clostridium difficile infectionClostridium difficile(CD)is a gram-positive,anaerobic and spore-forming pathogen.Since this opportunistic pathogen is confirmed to be associated with pseudomembranous colitis which is induced by long-term use of antibiotic in 1978,Clostridium difficile has become one of the most common pathogenic causes of hospital-acquired diarrhea.In Europe and North America,the morbidity of Clostridium difficile infection(CDI)has exceeded that of methicillin-resistant Staphylococcus aureus.With the abuse of antibiotics,especially broad-spectrum antibiotics,and the outbreak of highly virulent strains in western developed country,the morbidity,recurrence rate,and mortality of CDI have increased dramatically worldwide.In China,only a few hospitals have gradually carried out clinical monitoring of CDI,but the incidence of missed diagnosis and misdiagnosis of CDI by clinicians is still relatively high,and little is known about the risk factors for severe hospital-acquired CDI and the effect of different types of Clostridium difficile on the prognosis of patients.From September 2009 to September 2016,We had monitored hospital-acquired CDI in a tertiary teaching hospital.A total of 307 hospital-acquired CDI patients were enrolled.Thirty-three different sequence types(STs)were identified,among which ST-54(56/307,18.2%),ST-35(41/307,16.6%)and ST-37(46/307,15.0%)were the most prevalent.During the follow-up period,66 patients developed severe CDI and 32 patients died in 30 days.Multivariate logistic analysis revealed that bloodstream infection,pulmonary infection and C-reactive protein were significantly associated with severe CDI.After adjustment for potential confounders,old age,bloodstream infection,fever,mechanical ventilation,connective tissue disease,macrolide use and hypoalbuminaemia were independently associated with 30-day mortality in patients with hospital-acquired CDI.In addition,the prognosis of patients in the present study was closely related to toxin types and STs of Clostridium difficile.Our data can provide effective support for clinician to find high-risk patients,improve early diagnosis rate and infection control,and find effective prevention and treatment strategies for CDI.Part 2 Protective Effect of Pediococcus pentosaceus LI05 against Clostridium difficile infection in a mouse modelThe use of antibiotics can disrupt the structure of intestinal flora,and reduce species diversity and richness,then cause Clostridium difficile-associated diarrhea.At present,the clinical treatment of CDI is mainly the use of metronidazole and vancomycin.However,more than 20%of patients with CDI have experienced recurrence,costing more than $1.5 billion in the United States annually.Studies have provided evidence tor probiotics could be instrulental in restoring the intestinal dysbiosis caused by CDI.Here,we examined the protective effect of Pediococcus pentosaceus LI05 in a mouse CDI model.C57BL/6 mice were administrated P.pentosaceus LI05(LI05 group)or sterile anaerobic PBS(CDI group)everyday for 14 days.Mice were exposed to antibiotics cocktail and then challenged with Clostridium difficile strain VPI10463.Mice were monitored daily for survival and weight loss.Colonic tissue and serum samples were assessed for intestinal histopathology,intestinal barrier function and systemic inflammation.Fecal microbiome was analyzed by 16S rRNA sequencing and metabolome was detected by gas chromatography-mass spectrometry(GC-MS).The oral administration of P.pentosaceus LI05 improved the survival rate and alleviated the histopathological impact of Clostridium difficile.Compared to the CDI group,the levels of inflammatory mediators in the colon as well as inflammatory cytokines and chemokines in serum were substantially attenuated in the LI05 group.P.pentosaceus LI05 alleviated the CDI-induced of disruption of ZO-1,occludin and claudin-1,and Improve intestinal metabolites.Additionally,fecal microbiome analysis showed an enrichment in the abundance of Rikenellaceae,Porphyromonadaceae and Deferribacteraceae,while,the relative abundance of Escherichia/Shigella were decreased.Our results demonstrated that the preventive effect of P.pentosaceus LI05 against CDI was mediated via improving intestinal barrier function,down-regulating the inflammatory response,regulating intestinal flora and fecal metabolome.Therefore,P.pentosaceus LI05 could be a promising probiotic in CDI.