The Association Of Gut Microbiome And Its Metabolism With Excessive Gestational Weight Gain In Early Pregnancy

Objectives（1）The 16S ribosomal RNA gene amplicon analysis was applied to discuss the association between gut microbiome and EGWG in the first trimester.（2）The untargeted metabolomics analysis was applied to assess the association of stool and plasma metabolitic profile with EGWG in the first trimester.（3）The results of 16S rRNA amplicon analysis and untargeted metabolomics analysis were integrated to explore the host-gut microbiome co-metabolism pathway associated with EGWG.Methods（1）This study was a nested case-control study based on a cohort established in the first trimester(8～13⁺⁶weeks).The EGWG and AGWG groups were identified via Chinese diagnostic criteria:Weight monitoring and evaluation during pregnancy period of Chinese women（T/CNSS009-2021）.Participants were screened based on inclusion and exclusion criteria;some of them were further excluded because of lacking stool or blood samples.A total of 50 EGWG pregnant women were randomized sampled via random number table;another comparable 50 AGWG counterparts were included in this study.（2）The 16S ribosomal RNA gene amplicon analysis was applied to test fecal samples in order to explore the characteristic of microbiome structure,the association between microbiome and the influence factors,the interaction among genuses,as well as to identify the different micriobiome and predicted function modules between groups.（3）Liquid Chromatograph Mass Spectrometer was applied in the untargeted metabolomics analysis to explore the character of metabolites and the different metabolites and metabolic pathways between groups.（4）The metabolome-microbiome-metadata correlation analysis（3MCor）was applied to explore the association of identified genuses and metabolites in global,hierarchical and pairwise level.Results（1）The study cohort enrolled 713 participants at the beginning.A total of 601 participants with single birth were recorded.According to the Chinese diagnostic criteria,260 women were EGWG,301 women were AGWG,and 40 women were inadequate gestational weight gain.The incidence of EGWG in singleton pregnancy women was 43.3%（260/601）.16S ribosomal RNA gene amplicon analysis indicated that the most abundant genuses were Faecalibacterium,Blautia,Bacteroides,Coprococcus,and Roseburia.The ratio of Prevotella/Bacteroides in EGWG was higher than AGWG（P<0.05）.No significant Alpha diversity was found.Significant Beta diversity was identified by permutational multivariate analysis of variance（between group q value=0.017）,non-metric multidimensional scaling（stress=0.174）and partial least squares discriminant analysis.A total of 13 different genuses between groups were identified,including Alistipes,Bacteroides,[Prevotella]（Paraprevotellaceae famliy）,Butyricimonas,Stenotrophomonas,Pseudomonas,Serratia,Enterobacter,Ruminococcus,Anaerofustis,Abiotrophia,Staphylococcus and Prevotella.There were various associations between family,genuses and environmental factors.There were extensive and complex associations between the genera.A total of14 predicted function modules were identified discrepantly between groups,including glycosaminoglycan degradation,glycosphingolipid biosynthesis-ganglio series,glycosphingolipid biosynthesis-ganglio series,glycosphingolipid biosynthesis-globo and isoglobo series,secondary bile acid biosynthesis,D-glutamine and D-glutamate metabolism,various types of N-glycan biosynthesis,sphingolipid metabolism,lysosome,ferroptosis,synthesis and degradation of ketone bodies,biofilm formation-Escherichia coli,pantothenate and Co A biosynthesis and C5-branched dibasic acid metabolism.（2）The untargeted metabolomics analysis indicated that metabolite of fecal and plasma did not separate obviously between two groups on a whole.A total of 15 fecal metabolite and 3 plasma metabolite were 2 fold change between groups（P<0.05）in univarite analysis.A total of 77 fecal metabolite and 29 plasma metabolite were VIP>1（P<0.05）identified via the orthogonal projections to latent structures discrimination analysis.Thirteen metabolic pathways with 9 different metabolites between two groups were identified by fecal samples.These thirteen metabolic pathways included biotin metabolism,pantothenate and Co A biosynthesis,lysine biosynthesis,lysine degradation,pyrimidine metabolism,tryptophan metabolism,porphyrin and chlorophyll metabolism,synthesis and degradation of ketone bodies,phenylalanine,tyrosine and tryptophan biosynthesis,propanoate metabolism,valine,leucine and isoleucine degradation,butanoate metabolism,tyrosine metabolism.These nine metabolite included biotin,pantothenate,L-Saccharopine,saccharopine,3-Dehydroquinate,acetoacetate,indoleacetate,bilirubin,and cytosine.The synthesis and degradation of ketone bodies pathway and acetoacetate were the most important（impact=0.250,P=0.015）.No different metabolic pathway between groups was identified by plasma samples.（3）3Mcor indicated that identified genuses were associated respectively with metabolites of fecal and plasma in global level.Dimensionality reduction was applied by weighted gene association network analysis.Three gut microbiome modules,10 fecal metabolite modules and 7 plasma metabolite modules were identified after filtered by phenotype with P value of 0.05.After adjusted for potential influence facoters,5 pairs of statistically significant associations were identified among 3 gut microbiome modules and 4 fecal metabolites modules while2 pairs of statistically significant associations were identified among 1 gut microbiome module and 2 plasma metabolites modules.The pairwise analysis between identied genuses and metabolites indicated that Anaerofustis,Prevotella,Stenotrophomonas,[Prevotella]（Paraprevotellaceae family）,Alistipes and Bacteroides were associated with various fecal metabolites while[Prevotella]（Paraprevotellaceae family）and Alistipes were associated with various plasma metabolites.Conclusions（1）The characteristic of gut microbiome associated with EGWG in the first trimester.No significant difference of Alpha diversity was found while significant difference of Beta diversity was identified between EGWG and AGWG group.A total of different 13 genuses and 14predicted function module were identified between groups.（2）The stool and plasma metabolitic profile associated with EGWG in the first trimester.Metabolites of fecal and plasma did not separate obviously between two groups on a whole.A total of 77 fecal metabolite and 29 plasma metabolite were VIP>1（P<0.05）.Thirteen metabolic pathways with 9 different metabolites between two groups were identified by fecal samples,among which the synthesis and degradation of ketone bodies pathway and acetoacetate were the most important.Acetoacetate may be of significant importance in gestational weight gain.（3）The gut microbiome and its metabolite influenced EGWG by multiple pathways in the first trimester.In combination with the characteristics and the predicted functional module of gut microbiome,differential metabolites and enrichment metabolic pathways,it was found that gut microbiome and its metabolites influenced the gestational weight gain through the mechanism of influencing the level of inflammatory response,insulin resistance and blood glucose,fat metabolism,endocrine function and so on.（4）The gut microbiome and its metabolite influenced EGWG in the first trimester.The characteristic gut microbiome and metabolite associated in global,hierarchical and pairwise.Alistipes,[Prevotella]（Paraprevotellaceae family）and Stenotrophomonas were associated with various metabolites.Several unreported associations between characteristic gut microbiome and metabolites were identified;and the mechanisms and the effects on gestational weight gain need further research.