Study On The Difference Of Intestinal Flora In Children With Normal Weight,Overweight And Obesity

This study intends to use high-throughput sequencing technology to sequence 16 SrDNA of intestinal microflora in children with different body weights,and to compare the species diversity and community composition of the microbial communities;quantitative Real-time PCR was used to detect Eubacterium rectale,Bacteroides thetaiotaomieron,Bifidobacterium and Lactobacillus in children with different body weights,and their quantitative differences were compared.According to the study,we should establish a comprehensive understanding of the intestinal microecological structure of obese children,analyze the characteristics of intestinal micro-ecological imbalance in obese children,explore the relationship between intestinal flora and obesity,and provide a theoretical basis for the prevention and control of obesity by regulating the intestinal microecology.Objective:1.The diversity and community composition of intestinal microflora in normal weight,overweight and obese children were compared by high throughput sequencing technology,and to analyze the differences in intestinal flora between children with different body weights.2.Quantitative Real-time PCR was used to detect the number of Eubacterium rectale,Bacteroides thetaiotaomieron,Bifidobacterium and Lactobacillus in normal weight,overweight and obese children,and to analyze the differences in intestinal flora between children with different body weights.Methods:1.A total of 100 feces were collected from 30 normal weight,35 overweight and 35 obese children.Genomic DNA of the samples were extracted for amplifying the 16 S rDNA V4 hypervariable region by PCR and High-throughput sequencing by Illumina-HiSeq.The composition,diversity and richness of microbial communities were compared among the three groups.2.A total of 166 feces were collected from 54 normal weight,53 overweight and 59 obese children.Genomic DNA of the samples were extracted from stool,bacterial copy numbers were quantified by quantitative Real-time PCR with primers specifically targeting V3 region of the 16 S r DNA using the StepOne Real-Time PCR system.Kruskal-Wallis H test was used to compare the difference between groups,P-values<0.05 was considered statistically significant.Result:1.With Illumina sequencing,we determined a total of 4320570 clean 16 S rDNA V4 tags.There was no significant difference in Shannon index and Simpson index among the three groups.But a significant difference was found in observed_species,Chao1 index and PD whole tree.Beta diversity analysis showed that there was significant difference in the composition of intestinal community between normal weight and obese children(P < 0.05),while the composition of intestinal community of overweight was similar to normal weight and obese children.We further used LEfSe to evaluate the bacterial groups with significantly different percentages among normal weight,overweight and obese children.Bacteroides uniformis,S24-7,Holdemania,Guilliermondii,Christensenella,Peptostreptococcaceae,Megasphaera,Clostridium,Clostridium hathewayi,Anaerotruncus Iens,Bifidobacterium.adolescentis,Oxalobacteraceae,Formigenes,Deltaproteobacteria,Desulfovibrionales,Desulfovibrionaceae,Desulfovibrio were the biomarkers of the normal weight group.[Prevotella] and Christensenellaceae increased significantly in overweight group.Proteobacteria,Betaproteobacteria,Burkholderiales,Alcaligenaceae,Sutterella,Pasteurellales,Pasteurellaceae,Aggregatibacter,Haemophilus,Parainfluenzae,Prevotellaceae,Prevotella,shrinking,shrinking Acteriaceae,Fusobacterium,Leptotrichiaceae,Sneathia,Bacteroides coprophilus,Bacteroides.eggerthii,Veillonella,Veillonella.dispar,Megamonas,Ruminococcus gnavus were significantly increased in obesity group.2.The Eubacterium rectale,Bacteroides thetaiotaomieron,Bifidobacterium and Lactobacillus were significant difference among three groups.The median number of Eubacterium rectale in the normal weight,overweight,and obesity groups were 6.50(QR 5.83-7.02),6.79(QR 6.28-7.38),and 7.34(QR 6.91-7.78),respectively.The median numbers of Bacteroides thetaiotaomieron in the normal weight,overweight,and obesity groups were 5.00(QR 4.93-5.74),4.77(4.69-5.44),and 4.79(QR 4.69-5.36),respectively.The median numbers of Bifidobacterium in the normal weight,overweight,and obesity groups were 8.31(QR 8.18-8.54),8.31(QR 8.11-8.47),and 7.33(QR 7.16-7.57),respectively.The median numbers of lactobacillus in the normal weight,overweight and obesity groups were 6.12(QR5.90-6.32),5.85(QR 5.70-5.97)and 5.29(QR 5.01-5.63),respectively.Conclusion:1.With the increase of BMI,there is a decreasing tread in the amount of species abundance.There was no significant difference in the alpha diversity among the three groups.Beta diversity analysis showed that there was significant difference in the composition of intestinal community between normal weight and obese children(P < 0.05),while the composition of intestinal community of overweight was similar to normal weight and obese children.The proportion of some specific bacteria in the three groups has changed.2.The increase of Eubacterium rectale may be a stimulative effect of overweight and obesity.however,Bacteroides thetaiotaomieron,Bifidobacterium and Lactobacillus have a potential opposite effect.The number of Eubacterium rectale,Bacteroides thetaiotaomieron,Bifidobacterium and Lactobacillus in the intestine of normal weight,overweight and obese children was less affected by children's gender and age.