Oral <i>Lactobacillus reuteri</i> Combined With Intravenous Lysis‐Circuit‐Bearing Engineered Bacteria RE Enhances Antitumor Efficacy

Engineered bacteria can preferentially colonize hypoxic tumors, but improving efficacy while limiting systemic bacterial burden remains a central challenge. Here, we combined oral Lactobacillus reuteri (LR) with intravenous administration of RE, an engineered Escherichia coli Rosetta strain carrying the quorum-sensing lysis plasmid pTD103-E. This circuit couples LuxI/R-mediated population sensing to expression of the phage lysis gene φX174 E and was designed to enable density-associated bacterial lysis. In a 4T1 mouse model, LR+RE improved antitumor efficacy by 45% compared with single-bacterium treatment. RE remained enriched in tumors while being cleared from major organs within 72 h. Combined treatment increased intratumoral IFN-γ, altered IL-12 expression, elevated AhR abundance in tumor tissues, and increased CD8+ T-cell infiltration. These changes were accompanied by polarization of tumor-associated macrophages toward an M1-like phenotype, suppression of M2-like features, and increased markers of cell-cycle arrest and apoptosis. Fecal 16S rRNA profiling further showed that treatment reshaped the gut microbiota and identified Prevotellaceae_NK3B31_group as a microbial taxon associated with IFN-γ-centered antitumor responses. Together, these findings indicate that a natural probiotic can potentiate a tumor-targeted engineered bacterial therapy and is associated with coordinated microbiota-immune remodeling.