Supplementary Material for: Hydration Biomarkers Are Related to the Differential Abundance of Fecal Microbiota and Plasma Lipopolysaccharide-Binding Protein in Adults

<b><i>Introduction:</i></b> Prevalence of chronic hypohydration remains elevated among adults in the USA; however, the health effects of hypohydration in regards to human gut health have not been explored. <b><i>Methods:</i></b> This study examined the relationship between total water intake, hydration biomarkers (first-morning urine specific gravity [FMU_sg], first-morning urine volume [FMU_vol], and plasma copeptin), fecal microbiota, and plasma lipopolysaccharide-binding protein (LBP) in adults (25–45 years, 64% female). Fecal microbiota composition was assessed using 16S rRNA gene sequencing (V4 region). Immunoassays quantified plasma copeptin and LBP in fasted venous blood samples. Dietary variables were measured using 7-day food records. Linear discriminant analysis effect size (LEfSe) analyzed differentially abundant microbiota based on median cutoffs for hydration markers. Multiple linear regressions examined the relationship between LBP and copeptin. <b><i>Results:</i></b> LEfSe identified 6 common taxa at the genus or species level that were differentially abundant in FMU_sg, total water (g/day), or plasma copeptin (µg/mL) groups when split by their median values. Uncultured species in the <i>Bacteroides</i>, <i>Desulfovibrio</i>, <i>Roseburia</i>, <i>Peptococcus</i>, and <i>Akkermansia</i> genera were more abundant in groups that might indicate poorer hydration status. Multivariate linear analyses revealed a positive relationship between plasma copeptin and LBP when controlling confounding variables (<i>F</i>(6,52) = 4.45, <i>p</i> = 0.002, <i>R</i>² = 0.34). <b><i>Conclusions:</i></b> Taxa common between markers are associated with the intestinal mucus layer, which suggests a potential link between hydration status and intestinal mucus homeostasis. The relationship between LBP and copeptin indicates that copeptin may be sensitive to metabolic endotoxemia and potentially gut barrier function.