Engineered Bacillus subtilis as oral probiotics to target circulating lactic acid

Abstract

Elevated lactate concentrations are implicated in various acute and chronic diseases such as sepsis and mitochondrial dysfunction, respectively. Conversely, ineffective lactate clearance is associated with poor clinical prognoses and high mortality in these diseases. While several groups have proposed using small molecule inhibitors and enzyme replacement to reduce circulating lactate, there are few practical and effective ways to manage this condition. Recent evidence suggests that lactate is exchanged between systemic circulation and the gut, allowing bidirectional modulation between the gut microbiota and peripheral tissues. Inspired by these findings, this work seeks to engineer spore-forming probiotic B. subtilis strains to enable intestinal delivery of lactate oxidase as a therapeutic enzyme. After strain optimization, we showed that oral administration of engineered B. subtilis spores to the gut of mice reduced elevations in blood lactate in two different mouse models involving exogenous challenge or pharmacologic perturbation without disrupting gut microbiota composition, liver function, or immune homeostasis. Taken together, through the oral delivery of engineered probiotic spores to the gastrointestinal tract, our proof-of-concept study offers a practical strategy to aid in the management of disease states with elevated blood lactate and provides a new approach to knocking down circulating metabolites to help understand their roles in host physiological and pathological processes. Significance StatementThis study pioneers the use of engineered Bacillus subtilis spores as an oral probiotic therapy to enhance the clearance of elevated blood lactate, a condition linked to severe health issues like sepsis and metabolic disorders. By genetically modifying these spores to deliver therapeutic enzymes directly to the gut, we demonstrated a practical, effective method to modulate systemic lactate levels. This approach leverages the natural exchange between the gut microbiota and systemic circulation, offering a new strategy for managing diseases associated with lactate dysregulation. The safety and efficacy of this method were validated in mouse models, providing a foundation for future clinical applications.