The Study Of Correlation Between Gut Microbiota And The Traditional Chinese Medicine Syndrome Types Of Functional Constipation In Children

Objective:Based on the literature review of the research progress on the correlation between intestinal microbiota and the traditional Chinese syndrome（TCM）types of Functional Constipation（FC）in children,the clinical information and TCM syndrome type of pediatric FC were studied by statistical method.The 16SrDNA technology was used to compare and analyze the differences of intestinal flora between children with FC and healthy children with different syndrome types,genders,delivery modes and feeding modes,and to find the specific strains of FC in children with different syndrome types,thus providing a new direction for clinical diagnosis and treatment of FC and demonstrating the scientific nature of TCM syndrome differentiation and treatment from the aspect of intestinal flora.Methods:1.Clinical study:（1）A total of 45 children with FC were collected according to the inclusion criteria and exclusion criteria,and their basic information,general situation and four-diagnosis information were recorded.The unified constipation severity score and TCM syndrome score scale were filled in,and the syndrome differentiation was conducted.Afterwards,the collected information and syndrome scores were imported into Excel.（2）The distribution of FC children with different age,gender,delivery mode,feeding mode and the correlation between age,gender,delivery mode,feeding mode,height,weight and constipation severity were analyzed.（3）Based on SPSS statistical software,cluster analysis was conducted on the information of the four diagnoses of FC in children,and the distribution characteristics of the information of the four diagnoses were obtained.（4）Combined with the information distribution characteristics of the four diagnoses and TCM syndrome typing standards,the distribution characteristics of TCM syndrome types of pediatric FC in this study were studied,and the correlation between the distribution characteristics and age,gender,delivery mode,feeding mode,etc.2.Experiment research:16SrDNA technology was used to detect intestinal flora in children with FC of different syndromes and healthy children.To analyze the differences of intestinal flora between children with FC and healthy children,the differences of intestinal flora among children with different genders,delivery modes and feeding modes who suffered from FC,the characteristics of intestinal flora in children who occurred FC with different syndromes,and the differences between intestinal flora in children with typical symptoms and children without typical symptoms.Results:1.A total of 45 children with FC were collected in this study,including 18 males（40%of the total）and 27 females（60%of the total）,with a male to female ratio of 2:3.The minimum age was 2 years old,the maximum age was 14 years old,with an average age of 5.69±2.512 years old.There were 5 cases in the infant group,25 cases in the pre-school age group,11 cases in the school-age group,and 4 cases in the adolescent group.There was no statistical difference in gender distribution among different age groups and no statistical difference in age distribution among different gender groups（P>0.05）.There was no statistical difference in the proportion of children with FC by gestational age at birth,feeding mode,time of stopping breast milk and time of supplementary food addition（P>0.05）.There was no statistical difference in the severity of constipation in children with FC by gender,age,feeding mode,time of breast-feeding and supplementary feeding.There was no significant correlation between the severity of constipation and body weight and height.There were significant differences in the severity of constipation in children with FC in different delivery modes（P=0.018<0.05）.The mean value of constipation severity in children with cesarean delivery was 12.31±4.750,and that in children with vaginal delivery was 8.44±3.926,indicating that the severity of constipation in children with cesarean delivery was higher than that in children with vaginal delivery.2.In the clinical judgment of TCM syndromes and cluster analysis of the information of the four diagnoses based on SPSS statistical software,it was found that most of the TCM syndromes of pediatric FC have not only a single syndrome,and the combined syndromes accounted for the majority.Among the combined syndrome,33.3%were spleen deficiency combined with food accumulation syndrome,24.4%were food accumulation combined with heat syndrome,24.4%were spleen deficiency syndrome,11.1%were food accumulation syndrome and 6.7%were heat syndrome.3.There was no significant difference in TCM syndrome type distribution of FC children with different age,gender,delivery mode and feeding mode（P>0.05）.4.There were no significant differences in a diversity index and β diversity index between children with FC and healthy children,children with different genders,different delivery modes,different feeding modes and different TCM syndrome type of FC（P>0.05）.5.The P values of Anosim analysis and Adonis analysis among children with FC and healthy children were all greater than 0.05,indicating that the differences between the two groups and intra-group differences were not statistically significant,and the intestinal microbes of the two groups could not be significantly distinguished.Metastats difference analysis showed that the dominant flora of the two groups could be obtained at the genus level.The dominant flora of the children with FC mostly belonged to firmicutes rumen bacteria family which is mainly composed of Eisenbergiella,Oscillospiraceae_Unclassified,UBA1819,Ruminococcaceae_Unclassified,Erysipelatoclostridium（P<0.05）.The dominant flora of healthy children were Veillonella,Haemophilus,Subdoligranulum and Lactobacillus（P<0.05）.6.Differences of intestinal flora in children with different genders,delivery modes and feeding modes:The P values of Anosim analysis and Adonis analysis among children with FC of different genders were all greater than 0.05,indicating that the difference between the two groups and the difference within the group were not statistically significant,and the intestinal microbes of the two groups could not be significantly differentiated.Metastats difference analysis found that at the genus level,the abundance of Lachnospira,Coriobacteriales_Incertae_Sedis_Unclassified and Ruminococcaceae_Unclassified in male group was higher than that in female group.The abundance of Lactobacillus in the girl group was higher than that in the boy group（P<0.05）.The P values of Anosim analysis and Adonis analysis among children with FC of different delivery modes were all greater than 0.05,indicating that the differences between the two groups and intra-group differences were not statistically significant,and the intestinal microbes of the two groups could not be significantly differentiated.Metastats analysis showed that Megamonas was higher in the vaginal delivery group than in the cesarean delivery group,and Lachnospira was higher in the cesarean delivery group than in the vaginal delivery group（P<0.05）.Analysis of differences between Anosim groups of FC children with different feeding methods showed that R=0.162>0,P=0.007<0.05,there was statistical difference among the three groups,and the difference between groups was greater than the difference within the group.Adonis analysis showed that the value of F in Fisher test was 1.793,R=0.098,P=0.005<0.01,indicating that the intestinal microflora of the three groups could be significantly distinguished,indicating that different feeding methods had an impact on the intestinal microflora.LefSe analysis found differences in phylum,class,order,family and genus levels among the three groups.Firmicutes were the dominant flora in the intestinal tract of children with FC who were exclusively breastfed,and actinobacteria were a small part.Firmicutes and proteobacteria were the dominant flora in mixed feeding and artificial feeding.Among them,the dominant flora in breast-feeding group were Firmicutes,Clostridia,Lachnospirales,Lachnospiraceae,Agathobacter.Order Oscillospirales,DTU089,Ruminococcus,Incertae Sedis,UCG 005,NK4A214 Group,Anaerofustaceae,Anaerofustis,Gordonibacter.The dominant bacteria in the mixed feeding group were Proteobacteria,Gammaproteobacteria,Enterobacterales,Enterobacteriaceae and Escherichia_Shigella,Turicibacter,Lachnoclostridium,Anaerovoracaceae.The dominant flora in artificial feeding group were Burkholderiales,Sutterellaceae,Christensenellaceae_R7group and[Eubacterium]_ventriosum_group.7.Differences of intestinal flora in children with FC of different TCM syndromes:The P values of Anosim analysis and Adonis analysis among children with FC of different TCM syndromes were>0.05,indicating that the differences between groups and intra-group differences were not statistically significant,and the intestinal microbes of each group could not be significantly differentiated.The intestinal microflora structure of children with FC of different TCM syndromes is different,and there is no dominant micro flora in the heat syndrome group and the food accumulation combined with heat syndrome group.The dominant flora of the food accumulation syndrome group was Synechococcales,Cyanobiaceae,Cyanobium_pCC₆307,and the dominant flora of the spleen deficiency combined with food accumulation syndrome group was Subdoligranulum.Negativicutes were the dominant flora in spleen deficiency syndrome group.Conclusion:1.The proportion of FC in children is the largest in pre-school age.There was a correlation between the severity of constipation and delivery mode,and the severity of constipation in cesarean delivery was higher than that in vaginal delivery.2.Children’s FC is mainly combined with syndrome,and the TCM syndrome mainly includes the following five types:spleen deficiency combined with syndrome,heat caused by accumulation of food,spleen deficiency combined with syndrome caused by accumulation of food,and heat caused by accumulation of spleen.3.Difference of intestinal flora between children with FC and healthy children:The dominant flora of children with FC is firmicutes rumen bacteria family.The dominant flora of children with FC were Eisenbergiella,Oscillospiraceae_Unclassified,UBA1819,Ruminococcaceae Unclassified,Erysipelatoclostridium（P<0.05）.The dominant flora of healthy children were Veillonella,Haemophilus,Subdoligranulum and Lactobacillus（P<0.05）.4.There were differences in intestinal microflora community structure in children with functional constipation by gender,delivery mode and feeding mode,and the dominant microflora existed at the genus level.5.Differences of intestinal flora in children with FC of different TCM syndromes:There is no dominant microflora in the heat syndrome group and the food accumulation combined with heat syndrome group.The dominant flora of the food accumulation syndrome group was Synechococcales,Cyanobiaceae,Cyanobium_pCC₆307,and the dominant flora of the spleen deficiency combined with food accumulation syndrome group was Subdoligranulum.Negativicutes were the dominant flora in spleen deficiency syndrome group.