Impact Of Antibiotic On Infants' Gut Microbiota By Culturomics And Metagenomics

The discovery and clinical use of antibiotics can effectively control bacterial infection.And antibiotics play an important role in the prevention and treatment of disease in infants and young children.However,the usage of antibiotics affects the normal intestinal flora,causing intestinal microecological disorder and intestinal diseases such as antibiotic associated diarrhea,inflammatory bowel disease,irritable bowel syndrome,etc.And it can induce metabolic,immune and neurodevelopmental disease such as obesity,diabetes,autism,type 2 allergies,asthma and anxiety.The use of antibiotics in early life has a long-term impact on the intestinal flora and the normal growth of body.Therefore,it is necessary to study the influence of antibiotics on the composition,diversity and function of intestinal flora in infants.As to provide scientific basis for the rational use of antibiotics in clinical practice and the recovery of intestinal flora.In this study,the intestinal flora of healthy infants and antibiotic intake infants was taken as the research object.Combined with culturomics and SMRT sequencing technology,the culturable microorganisms were isolated and identified.The diversity and composition of culturable microorganisms in the intestinal tract of infants were analyzed.Fecal samples were analyzed by metagenomics sequencing,which fully revealed the diversity of intestinal flora caused by antibiotics in infants and the differences in composition at different taxonomic levels.This study determined the influence of antibiotics on bacteria flora function and the pathways.And it revealed the influence on intestinal resistance genes.Through the above research,the results are as follows:(1)Based on culturomics and SMRT sequencing technology,the distribution of culturable microorganisms in infants' intestine and the influence of antibiotics on intestinal diversity were analyzed.Culturomics was used to separate the intestinal flora of healthy and amoxicillin intake infants.Then 6234 bacterial colonies were obtained.After 16 S r DNA and SMRT sequencing identification,all colonies could be divided into 3 phyla,16 genera and 45 species,among which Enterococcus(49.70%)and E.faecalis(32.91%)was the dominant genus and species in infants' intestinal flora.By alpha and beta diversity analyze,there were differences in the composition of culturable microorganisms between the antibiotic group and the healthy group.The Chao1 index,observed species index and Faith's PD index were significantly lower than that of healthy group(p(27)0.05).(2)The effect of antibiotics on the composition of culturable microorganisms in the intestinal flora were identified.According to taxonomic differences of intestinal culturable microorganisms,the relative abundance of Enterococcus was decreased by 6.89% and that of Klebsiella and Enterobacter increased by 20.62% and 6.60%,respectively,compared with healthy group.At species level,the relative of E.faecalis decreased by 20.88%.E.hormaechei and E.faecium increased by 7.33% and 19.18%,respectively.(3)The composition and distribution of intestinal flora in infants were analyzed to study the effects of antibiotics on the intestinal flora diversity and species classification.Metagenomic sequencing was used to study the taxonomic composition of intestinal flora.And dominant intestinal phyla,genus and species were identified as Actinobacteria(42.41%),Bifidobacterium(41.81%)and B.longum(32.03%).The results of beta diversity analysis showed that there were differences in the intestinal flora between the antibiotic group and the healthy group.The alpha diversity analysis found that the diversity index of the healthy group was higher than that of antibiotic group and the difference of Chao1 index was significant(p(27)0.05).Based on LEFSe analysis.It could be seen that the intake of antibiotics leads to the intestinal dominant bacteria and dominant species into Enterococcus(33.17%)and E.faecium(32.15%).Correlation analysis of dominant species revealed that C.sphenoides and R.mucilaginosa had strong correlation with other strains in the intestinal tract.(4)The effects of antibiotics on the function of microflora and the pathways were revealed.Based on the function prediction of KEGG protein database,the function of intestinal flora in infants was analyzed and it was found that the intake of antibiotics mainly affected the metabolic.Antibiotics enhanced carbohydrate metabolism of intestinal flora by inhibiting ethyl butyrate metabolic pathway and activating phosphate metabolic pathway.In addition,antibiotics also act on arginine and proline metabolism and lysine degradation to weaken amino acid metabolism.Based on Egg NOG protein database,antibiotics weaken the information storage of intestinal flora in infants and enhance the energy metabolism function of intestinal flora.Based on CAZy database,it was found that antibiotics attenuated the activity of PLs.GHs,CEs,AAs and enhanced the activity of GTs,CBMs.(5)It was clear that antibiotics cause increased resistance to intestinal flora.Based on ARDB antibiotic resistance annotation,the results found that drug-resistance gene in abundance in antibiotics group is higher than that in healthy group.All of the resistance gene expression for 11-alkenyl pyrophosphoric acid phosphatase and of bacitracin resistance.Most resistant gene were associated with K.pneumonia and related to the rate of 55.56%.