Comparative Analysis of Taxonomic and Functional Gut Microbiota Profiles in Relation to Seroconversion of Thyroid Peroxidase Antibodies in Euthyroid Participants

Background: Previous studies have reported gut microbiome alterations in Hashimoto’s autoimmune thyroiditis (HT) patients. Yet, it is unknown whether an aberrant microbiome is present before clinical disease onset in participants susceptible to HT or whether it reflects the effects of the disease itself. Here, we report for the first time a comprehensive characterization of the taxonomic and functional profiles of the gut microbiota in euthyroid seropositive and seronegative participants. Our primary goal was to determine taxonomic and functional signatures of the intestinal microbiota associated with serum thyroid peroxidase antibodies (TPOAb) antibodies. A secondary aim was to determine whether different ethnicities warrant distinct reference intervals for accurate interpretation of serum thyroid biomarkers. Methods: In this cross-sectional study, euthyroid participants with (N=159) and without (N=1,309) TPOAb were selected from the multi-ethnic (European Dutch, Moroccan, and Turkish) HEalthy Life In an Urban Setting (HELIUS) cohort. Fecal microbiota composition was profiled using 16S rRNA sequencing. Differences between the groups were analyzed based on overall composition (alpha and beta diversity), as well as differential abundance of microbial taxa and functional pathways using multiple differential abundance tools. Results: Overall composition showed a substantial overlap between the two groups (p>0.05 for alpha-diversity; p=0.39 for beta-diversity), indicating that TPOAb-seropositivity does not significantly differentiate gut microbiota composition and diversity. Interestingly, TPOAb-status accounted for only a minor fraction (0.07%) of microbiome variance (p=0.545). Further exploration of taxonomic differences identified 138 taxa nominally associated with TPOAb-status. Among these, thirteen taxa consistently demonstrated nominal significance across three additional differential abundance methods, alongside notable associations within various functional pathways. Furthermore, we showed that ethnicity-specific reference intervals for serum thyroid biomarkers are not required, as no significant disparities in serum thyroid markers were found among the three ethnic groups residing in an iodine-replete area (p>0.05 for TSH, fT4, and TPOAb). Conclusion: These findings suggest that there is no robust difference in gut microbiome between individuals with or without TPOAb in terms of alpha and beta-diversity. Nonetheless, several taxa were identified with nominal significance related to TPOAb presence. Further research is required to determine whether these changes indeed imply a higher risk of overt HT.