A host-directed adjuvant sensitizes intracellular bacterial persisters to antibiotics

Abstract Intracellular bacterial reservoirs contribute to antibiotic treatment failure by fostering metabolically dormant persister cells that are highly tolerant to killing. However, strategies to effectively target intracellular persister cells remain limited. Here we developed a high-throughput screen to identify compounds that modulate the metabolic activity of intracellular Staphylococcus aureus . The identified compound, KL1, increases intracellular bacterial metabolic activity and sensitizes persister populations of S. aureus to antibiotics, without causing cytotoxicity or bacterial outgrowth. KL1 also exhibits adjuvant activity against intramacrophage Salmonella enterica Typhimurium and Mycobacterium tuberculosis , as well as in murine infection models of S. aureus and S . Typhimurium infection. Transcriptomic analysis and further mechanistic studies reveal that KL1 modulates host immune response genes and suppresses the production of reactive species in host macrophages, alleviating a key inducer of antibiotic tolerance. Our findings highlight the potential to target intracellular persisters by stimulating their metabolism. There are two major problems in the field of antimicrobial chemotherapy–antibiotic resistance and antibiotic tolerance. Antibiotic tolerance has been frequently connected with poor treatment outcomes in the clinic. Unlike antibiotic resistance, which permits bacterial growth in the presence of drugs, antibiotic tolerance allows bacteria to withstand multiple antibiotics for prolonged periods. The extended survival of tolerant bacteria further predisposes them to evolve antibiotic resistance over time, underscoring the critical need to address antibiotic tolerance. Host interactions have been shown to induce persister formation in numerous pathogens, with the production of reactive oxygen and nitrogen species heavily implicated in the collapse of bacterial metabolic activity and entry into an antibiotic-tolerant state. Yet, tools to study or target this process remain limited. Here we developed a high-throughput screen to identify compounds that modulate intracellular S. aureus metabolism, leading to the discovery of KL1, a host-directed compound that sensitizes persisters to antibiotic killing.