Abstract
<title>Abstract</title> <p> Purpose Although prior research has explored the association between gut microbiota (GM) and breast cancer (BC), the underlying causal mechanisms and mediating pathways remain insufficiently elucidated. To address this gap, a two-step Mendelian randomization (MR) approach was utilized to examine the causality between 473 GM and BC risk, alongside the mediating role of 731 immune cell (IC) types. Methods Two-sample MR analyses were conducted using publicly available Genome-Wide Association Study data to explore the causal relationships between GM and BC. Results Bidirectional MR analysis revealed that IC activity partially mediates the causality between GM and BC, with sensitivity analyses reinforcing the robustness of these associations. Notably, five taxa were identified as positively correlated with BC risk. Furthermore, 39 ICs characteristics were linked to BC, with 17 ICs demonstrating an increasing risk. Specifically, in the two-step MR analysis, both the <italic>Rhodanobacter</italic> (IVW: P = 0.0153) and CD25hi CD45RA+ CD4 not Treg %CD4+ (IVW: P = 0.0179) or CD25hi CD45RA+ CD4 not Treg %T cell (IVW: P = 0.0093) were found to be causally associated with an increased risk of BC.the causal effect of <italic>Rhodanobacter</italic> on BC was partially mediated by CD25hi CD45RA+ CD4 not Treg %CD4+ (IVW: P = 0.0043) and CD25hi CD45RA+ CD4 not Treg %T cell (IVW: P = 0.0116). These mediating effects accounted for 4.64% and 5.02% of the cumulative causal effects, respectively. Conclusion The findings provide genetically supported evidence linking GM, immune modulation, and BC etiology, with MR analysis offering robust validation of the tripartite interaction. </p>