Clinical Characteristics Of Children With Clostridium Difficile Infection And Establishment Of Murine Model

Clostridium difficile(CD),widely distributed in the natural environment,is a gram-positive anaerobic bacillus that belongs to the normal intestinal flora of the human body.CD is a opportunistic pathogen whose overgrowth cause an imbalance in the intestinal flora and release toxins that cause CD infection(CDI).CDI is the main reason of antibiotic-associated diarrhea(AAD),which has become the attention of the global public health problem and increased the economic burden of the patient’s family and society.CD spores can be spread through feces,body contact and air flow.However,due to children’s poor hand hygiene habits and the widespread use of broad-spectrum antibiotics,the incidence of CDI in children has been on the rise in recent years,which is manifested by increased recurrence rate,severity and mortality.Therefore,effective diagnosis and treatment measures are urgently needed.CDI may present with mild to moderate diarrhea,pseudomembranous enteritis,peritonitis,sepsis,and even toxic megacolon,and the culture of toxigenic strains and cytotoxin neutralization test were the standards for the diagnosis of CDI.At present,the treatments of CDI include antibiotics,fecal microbiota transplantation(FMT)and monoclonal antibody.To study the clinical characteristics of CDI and establish animal model that can simulate the clinical CD infection process of human is of great significance for studying the pathogenesis of CDI,the preparation of vaccines and the discovery of new therapeutic methods.In this study,the clinical characteristics of CDI in children will be analyzed in depth,and the CDI murine model will be established through the suspension of clinically isolated viable strains of CD and the induction of a variety of antibiotics.Clinical characteristics of children with clostridium difficile infectionObjective:In order to know the condition of children with CDI in the department of gastroenterology of Children’s Hospital affiliated to Shanghai Jiao Tong University,the clinical characteristics of CDI in children were discussed.Methods:Collecting the clinical information of 71 cases diagnosed with CDI in the department of gastroenterology.To explore the clinical characteristics of CDI in children through the analysis of the basic information,antibiotic exposure history,acid inhibitors use history,clinical manifestation,laboratory examination and radiographic results,colonoscopy,treatment methods,treatment effect and relapse etc.Results:Of the 71 children diagnosed with CDI,71%had a history of antibiotic exposure and 12.8%had a history of acid inhibitors use.The clinical manifestations of 83.1%of the children was diarrhea,bloody stool and abdominal pain,accounting for 32%and 31%respectively,and only 11.3%had abdominal distension.The mean value of C-reactive protein was(11.21±7.521)mg/L,the mean value of procalcitonin was(0.820±0.396)ng/ml,and the positive rate of CD toxin detection was 62%.CT imaging findings of the abdomen included 8 cases of intestinal wall thickening and 11cases of intestinal gas and fluid accumulation.The results of colonoscopy suggested 5cases of pseudomembranous enteritis and 24 cases of chronic inflammation.In the first round of antibiotic treatment,metronidazole alone,vancomycin alone or a combination were used.There was no statistical difference in the effective rate,but there was a statistical difference in the recurrence rate(P<0.05).The recurrence rate of vancomycin alone was lower.87%of the children got better after the first round of antibiotic treatment,and 31%of them relapsed,with an average interval of 2.36weeks.In the children of recurrent CDI and non-recurrent CDI,there was no statistical difference in gender,clinical manifestations,proton pump inhibitor use history,while there was a statistical difference in antibiotic exposure history,especially cephalosporin antibiotics(P<0.05).The effective rate of reantibiotic treatment was 78%,the recurrence rate was 14.3%,and the effective rate of FMT was100%.There was no significant adverse reaction in FMT treatment,and the difference of effective rate was statistically significant(P<0.05).Conclusions:Antibiotic exposure was a high risk factor for children with CDI,diarrhea was the most common clinical manifestation,hematochezia and abdominal pain followed,and abdominal distension was rare.In the laboratory examination,C-reactive protein and procalcitonin were generally increased,and the positive rate of CD toxin detection was 62%.Abdominal CT imaging showed intestinal wall thickening and intestinal cavity accumulation of gas and fluid,and colonoscopy typically showed pseudomembranous enteritis.Vancomycin alone had a lower recurrence rate in first-round antibiotic treatment.A history of antibiotic exposure,especially cephalosporin antibiotics,was a high risk factor for recurrent CDI in children.The effective rate of FMT in children with recurrent CDI was higher than that with antibiotics again.Establishment of murine model of clostridium difficile infectionObjective:Murine model of CDI induced by antibiotic exposure was established to provide evidence for a more effective understanding of the pathogenesis of CDI.Methods:Eighteen male C57BL/6 mice(8 weeks old,weight 18-22g)were divided into groups A,B and C.Group A was the control group(six),while group B and group C were the model group(six each).In the experiments of group B and group C,all the mice received a mixture of antibiotics(kanamycin 0.4mg/ml,gentamycin 0.035mg/ml,polymyxin 850U/ml,metronidazole 0.215mg/ml,vancomycin 0.045mg/ml)for 5 consecutive days.On the last day before infection,all mice in the experimental group were given intraperitoneal injection of lincomycin(10mg/kg),and mice in the control group were given 0.3ml intraperitoneal injection of normal saline,and then CD suspension(10~6CFU/ml and 10~8CFU/ml)were given gavage to group B and group C respectively to obtain the CDI murine model.The body weight change,diarrhea and systemic conditions of the mice were monitored after the murine model was obtained.Fecal samples were collected for quantitative PCR detection of toxin genes,and the length of the blind colon was measured and histopathological analysis was performed when necessary to evaluate the success of the model establishment.Results and conclusions:There was no significant difference in body weight between the model group and the control group.No diarrhea was observed in the mice,no obvious wet tail,hair loss and decreased mobility was observed,and no death was found.After the fecal samples were collected,no positive results were found in the detection of CD toxin gene by real-time fluorescence quantitative PCR.Compared with the control group,there was no significant difference in colonic length and the length of cecum increased in the model group.