| Objectives The research is aimed at describing the characteristics of gut microbiota during 0-3 months infants who are healthy newborns at full-term,and flora of colostrum and mature milk during lactation in Chinese mothers of vaginal delivery,exploring the factors affecting the establishment and development of the early infant gut flora,and the composition of the human milk microbiota.So as to develop intestinal probiotics and breast milk probiotics for the Chinese population,and provide theoretical foundation for health guidance.Method A cohort of 20 mother-infant pairs(who are healthy and were delivered by vaginal with full-term)born in Maternal and Child Hospital of Hubei Province between May to July in 2020 were included.Then,collecting samples,including(1)faeces samples: meconium(the first stool);faeces at 1 month of age;faeces at 3 months of age;postnatal faeces of mother.The groups are named in order: Meconium,Month-1,Month-3,MF.(2)human milk samples: colostrum(within the first 5 days after delivery),mature milk(during 4-6 weeks after delivery).16 S rRNA sequencing was used to analyze the composition structure of gut microbiota and breast milk microbiota.Meanwhile,the health condition of the cohort,as long with associated factors was recorded through follow-up visit and questionnaires.So as to explore the factors affecting the establishment of the infant's early gut microbiota and maternal factors affecting the composition structure of the milk microbiota,and the correlation between the gut microbiota,breast milk microbiota and mother-infant health was sought in a comprehensive manner and providing theoretical support for the potential value of probiotic development.Results 1.Species annotation analysis of among three groups of Meconium,Month-1 and Month-3 showed that Proteobacteria,Firmicutes,Actinobacteria,Bacteroide were the dominant phylum;the percentages of each group are: Meconium(39.1%,34.8%,13.0%,12.9%),Month-1(30.4%,35.0%,18.2%,16.4%),Month-3(33.4%,23.5%,23.0%,20.1%);Escherichia-Shigella,Bifidobacterium,Bacteroides,staphylococcus,Clostridium sensu stricto 1,Klebsiella are the dominant genus,ranged from 10% to 35% of the groups: Meconium(34.85%,14.71%,11.68%,10.94%,2.51%,13.28%,12.77%),Month-1(16.48%,19.39%,11.31%,1.43%,15.30%,13.28%,12.77%),Month-3(15.96%,22.34%,16.15%,0.23%,18.22%,16.90%,0.97%).2.Alpha diversity analysis revealed differences in species composition amon g groups of Meconium,Month-1 and Month-3.The Chao1 index tended to decrease as infants grow up;the Shannon and Simpson indices varied dynamically: the lowest in Meconium,the highest in Month-1,and it's intermediate in Month-3.3.Analysis of PCA/PCo A revealed that Meconium was significantly different to Month-1 and Month-3.Meta Stats analysis show Fusobacteria is the biomarker.LEf Se analysis screened biomarker are Clostridiumsensustricto1,Clostridiaceae1,Bacillus,Bacillales(with LDA values>4),and there is no significant difference between Month-1 and Month-3.4.Species annotation analysis of COS and MB showed that Firmicutes,Proteobacteria,Bacteroidetes,Actinobacteria,Fusobacteria,Cyanobacteria,Verrucomicrobia were the dominant phylum,the percentages in each group as follows,COS: 80.32%,7.86%,6.62%,4.51%,0.49%,0.04%,0.04%;MB: 65.13%,16.23%,12.89%,4.45%,1.03%,0.10%,0.14%.Staphylococcus,Streptococcus,Pantoea,Bacteroides,Gemella,Prevotella 9,Lactobacillus,Faecalibacterium,Rothia were the dominant genus,the percentages in each group as follows,COS: 67.25%,14.25%,0.06%,3.78%,2.87%,0.11%,0.15%,3.92%,0.6%;MB:22.76%,27.54%,14.15%,6.93%,5.09%,6.03%,5.32%,2.26%,2.27%.Alpha diversity index analysis showed that Chao 1,observed species and Shannon indices were higher in COS than MB.5.Analysis of PCA/PCo A revealed differences in species composition between COS and MB.Significant differences were found between two groups.Meta Stats analysis show Proteobacteria is the phyla with significant differences.LEf Se analysis show biomarkers including Firmicutes,Bacillales,Lactobacillales,Enterobacteriaceae,Enterobacteriales,Gammaproteobacteria,Pantoea Streptococcus/Streptococcaceae,Staphylocodcaceae/Staphylococcus,(with LDA values >2).Adonis/Anosim Analysis suggested a better grouping effect(R=0.40),with an observable statistical difference(p<0.001***)and a high degree of explanation for the difference(R2=0.815).6.RDA/CCA analysis of the Meconium suggested that TCB and Head Circumference were significantly correlated with the dominant phylum.Network analysis revealed that Citrobacter,Streptococcus,Klebsiella and head circumference showed significant positive correlations(P < 0.01**,rho > 0.3);Staphylococcus was synchronously associated with TCB(P < 0.05*,rho > 0.3)and Height(P < 0.05*,rho >-0.3).RDA/CCA analysis suggested a greater correlation between height,head circumference and the distribution of samples from Month-1: Actinobacteria,Bifidobacterium,Enterococcus(P < 0.05*,rho > 0.5),and Enterobacteriaceae;Ambiguous taxa,Bacteroides,[Ruminococcus] gnavus group(P < 0.05*,rho <-0.5)were significantly correlated with Height;Bacteroidetes(P < 0.05*,rho>0.5),Verrucomicrobia,Lactobacillus(P<0.05*,rho<-0.5)were significantly correlated with head circumference.7.Correlation analysis shows that Streptococcus,Klebsiella were significantly correlated with head circumference at birth(P < 0.05*,rho > 0.8);Staphylococcus was significantly correlated with TCB and Height at birth(P < 0.05*),positively correlated with TCB(rho > 0.8),Staphylococcus was significantly correlated with TCB at birth(P < 0.05*)and with Height(rho <-0.8).8.Correlation network analysis of human milk biomarker and related maternal factors revealed that maternal BMI(both pre-pregnancy BMI and full-term BMI)may significantly affect human milk microbiota including Catenibacterium,Bergeyella,Clostridiaceae 1,Klebsiella,Prevotellaceae Ga6A1 group,[Clostridium] innocuum group,Gammaproteobacteria,Acidimicrobiia(P<0.05*),Klebsiella and Bergeyella were significantly correlated with pre-pregnancy BMI and full-term BMI(p < 0.01**),Bacteroidales S24-7 group,Prevotella 9,Prevotellaceae,Lachnospiraceae NK4A136 group,Anaerostipes [Eubacterium] eligens group,[Eubacterium] rectale group were significantly correlated with age.9.Correlation network analysis of MF and related maternal factors revealed that Proteobacteria(P<0.05*,rho<-0.5),Verrucomicrobia,Akkermansia(P<0.05*,rho>0.5)were significantly correlated with age,[Eubacterium] hallii group(P<0.05**,rho>0.5)were significantly correlated with pre-pregnancy BMI and full-term BMI.Conclusions 1.Proteobacteria,Firmicutes,Escherichia-Shigella,Bifidobacterium constitute the dominant species of gut microbiota of infants aged 0-3 months,as the age increases,the diversity and richness of species is decreases,intestinal flora of infant is varied greatly in the first month and remains stable in the next two months.Fusobacteria,Bacillales and Clostridium sensu stricto 1 were biomarkers.2.Staphylococcus,Citrobacter,Bacteroides,Enterococcus of meconium were correlated with the degree of jaundice.Citrobacter,Streptococcus,Klebsiella,Actinobacteria,Bifidobacterium,Bacteroides,were associated about growth and development of infants aged 0-3 months;Staphylococcus may indicate movement development of infant.3.Firmicutes,Proteobacteria,Bacteroidetes,Staphylococcus,Streptococcus,constitute the dominant species of human milk microbiota.As lactation progresses,the degree of richness and diversity of species is decreased,biomarkers are Bacillales,Staphylococcus,Lactobacillales,Streptococcus.4.BMI and age of maternal both influence maternal postnatal intestinal flora and lactation flora. |
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