The Change Of Clostridium Dififcile In Feces Of Wheezing Children And Immunologic Mechanism

Objective Detect the content of Clostridium difficile by real-time fluorescencequantitative PCR methods to make sure there have intestinal flora change in humanfecal samples and validate the relationship between gut flora and Wheezing disease.Atthe same time,by using ELISA to interpret the change of the IL-2and IL-4.Methods The detected control group fecal sample and serum were collectedfrom hernia and other elective surgery patients in the Pediatric Surgery,68cases werecollected.experimental cases were diagnosed as bronchiolitis or asthma and bothduring acute exacerbation of wheezing,collected from the department of the firstpediatrics respiratory medicine institute of pediatrics, First hospital of Jilin universityduring October in2013to October in2014.20cases of asthma and66caces ofbronchiolitis.Total DNA of bacteria was extracted from children’s fecal samples byKit.We designed primers based on the16S ribosomal RNA (16S rRNA) whichpossessed specialities of bacteria as amplified region.The amplified gene fragment of16S rRNA acquired from conventional PCR were used as standard.Analyzed thestander and sample by SYBR Green Ⅰ dye method,we can achive the stander culveand quantify the Clostrid ium difficile in fecal sample with the real time PCR methodthen observe the contents changes of Clostridium difficile in the intestinal ofwheezing children to understand the relation between the flora and the Clostridiumdifficile.Statistical analysis the change by GraphPad Prism5. Detecting the content ofIL-2and IL-4from the serum between experimental group and control groupaccording to ELISA kit,elucidate the change of Th1/Th2,Statistical analysis by curveexpert1.4, verify the immunologic mechanism of wheezing disease.Result1. Real-time fluorescent quantitative PCR can detect the Clostridium difficile inhuman fecal.2.The Clostridium difficile in bronchiolits childre higher than controlgroup,which have statistically significant, p＝0.0349.3.There have obvious statistically significant of the Clostridium difficile between asthma and control group,p＝0.044.4.The quantity of the Clostridium difficile has no significant changes frombronchiolits children to asthma children, p＝0.7219.5.The level of IL-2in bronchiolits and asthma children have statisticallysignificant compared with control group, p＝0.0224.6.The content of IL-4higher in bronchiolits and asthma children than controlgroup, p＝0.0373.Conclusion1.Real-time fluorescent quantitative PCR method we used is sensitivity andrepeatability,which was suitable for detecting the Clostridium difficile in human fecal.2.The content of the Clostridium difficiletistical was obvious increased both inbronchiolitis and asthma children. Indicate that the increase of Clostridium difficilecan be the cause of wheezing.3.The Clostridium difficile have no statistically significant among bronchiolitisand asthma, speculate the two is the different stage of one disease.4.There have a higher concentration of IL-2and IL-4in serum of wheezingchildren,so there have the same change.