In Situ Bioorthogonal Conjugation of Delivered Bacteria with Gut Inhabitants for Enhancing Probiotics Colonization

Clinical treatment efficacy of oral bacterial therapy has been largely limited by insufficient gut retention of probiotics. Here, we developed a bioorthogonal-mediated bacterial delivery strategy for enhancing probiotics colonization by modulating bacterial adhesion between probiotics and gut inhabitants. Metabolic amino acid engineering was applied to metabolically incorporate azido-decorated d-alanine into peptidoglycans of gut inhabitants, which could enable in situ bioorthogonal conjugation with dibenzocyclooctyne (DBCO)-modified probiotics. Both in vitro and in vivo studies demonstrated that the occurrence of the bioorthogonal reaction between azido- and DBCO-modified bacteria could result in obvious bacterial adhesion even in a complex physiological environment. DBCO-modified Clostridium butyricum (C. butyricum) also showed more efficient reservation in the gut and led to obvious disease relief in dextran sodium sulfate-induced colitis mice. This strategy highlights metabolically modified gut inhabitants as artificial reaction sites to bind with DBCO-decorated probiotics via bioorthogonal reactions, which shows great potential for enhancing bacterial colonization.