Fig. 1

Study design and analytical framework. The study was based on the Tongji-Huaxi-Shuangliu Birth Cohort, with data collected during early and middle pregnancy. Participants included GDM cases and controls matched in a 1:2 ratio for fatty acid assessments, and in a 1:1 ratio for 16S rRNA gene sequencing and shotgun metagenomic sequencing. Data acquisition encompassed demographic and clinical information through questionnaires, along with plasma and fecal samples for biochemical and microbiota analyses. Statistical analyses included conditional logistic regression, linear mixed-effects models, and Mendelian randomization, with mediation and interaction analyses to assess the role of microbiota. GDM, gestational diabetes mellitus; OGTT, 75-g oral glucose tolerance test; SNPs, single-nucleotide polymorphisms