Construction of an engineered Escherichia coli strain with enhanced intestinal colonization and anti-inflammatory efficacy in colitis.

Engineered probiotics are considered effective and safe therapeutic strategies for the treatment of various diseases. Escherichia coli Nissle 1917 (EcN) has been widely used as a chassis strain because of its safety and well-established genetic manipulation system. However, the limited intestinal colonization ability of EcN limits its potential as a chassis for the construction of synthetic probiotics. Here, an engineered EcN strain (EcN-CPM) with enhanced gastric acid and bile salt tolerance and improved intestinal adhesion was constructed. Oral administration of EcN-CPM to mice with colitis alleviated disease severity and reshaped the disordered gut microbiome by decreasing the abundance of Escherichia-Shigella while increasing the abundance of norank_f_Muribaculaceae. Mechanistically, the EcN-CPM supernatant directly promoted the proliferation of norank_f_Muribaculaceae, a short-chain fatty acid (SCFA)-producing genus. Targeted metabolomics revealed that EcN-CPM restored the DSS-induced depletion of SCFAs, which were negatively correlated with the abundance of Escherichia-Shigella and positively correlated with the abundance of norank_f_Muribaculaceae. Consistent with these findings, EcN-CPM treatment upregulated the expression of Treg-associated markers (Foxp3, Ctla4, and Cd25) and downregulated the expression of Th17-related genes (IL-17A and Roryt) in colonic tissues, restoring the IL-17A/Foxp3 ratio to homeostasis. Untargeted metabolomics further demonstrated that EcN-CPM uniquely restored the levels of seven anti-inflammatory metabolites depleted by DSS treatment. Collectively, these findings demonstrate that EcN-CPM alleviates intestinal inflammation by remodeling the gut microbiota to increase the production of SCFAs and anti-inflammatory metabolites, thereby driving a shift in Th17- and Treg-associated transcriptional signatures. This study establishes a platform for precision-designed synthetic probiotics with enhanced probiotic properties.