Recording the role of persisters in infection relapse

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PROJECT SUMMARY Beside the well‐researched antibiotic resistance, bacterial pathogens can survive antibiotic exposure through drug persistence. Persistence has been more difficult to study because of the transiency of the phenomenon, where some bacteria arrest growth, thus surviving several antibiotics. Resumption of growth by persister is thought to account for infection relapses but this has never been directly tested or demonstrated. With this proposal, we aim at answering the fundamental question: are persisters the future “relapsors” and thereby constitute a real reservoir for relapse of infections? In this project, we will make use of a remarkably useful model intracellular pathogen, Salmonella enterica serovar Typhimurium to establish whether Salmonella persisters are directly responsible for infection relapse. This proposal leverages further the development by our lab of model systems and tools to track persisters in the context of infection. First, in Aim 1 of this project, we will adapt a CRISPR‐based recorder that we recently built in the lab to record in the bacterial genome of persisters the state of growth‐arrest, thus leaving an indelible genetic scar in CRISPR arrays that is perpetuated and detectable in their progeny. We will further characterize this tool during infection of primary murine macrophages to determine if, in addition to recording growth arrest, it can also be used as recorder of length of growth arrest. We will also adapt pSCRATCH to record other signals in addition to growth arrest. Then, in Aim 2, we will use the growth‐arrest recorder to assess for Salmonella, for the first time, whether persisters are the direct reservoir for bacterial regrowth during relapse. This Aim will enable us to determine where relapsors originate from, from growth‐arrested persisters or few growers not eradicated by the drugs for other reasons, such as lack of antibiotic accessibility. It is of the utmost importance to determine whether persisters are the major source of infecting bacteria during relapse in order to efficiently target persistent infections with improved therapeutics and counteract the generation of antibiotic resistance.