Escaping the Salmonella-containing vacuole – Role of flagella-mediated damage, cytosolic motility, and host cell apoptosis in exit and transmission of Salmonella

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The project investigates the role of flagella and intracellular motility for the exit from the host cell. We previously observed that integrity of the Salmonella-containing vacuole (SCV) is decisive for the fate of intracellular Salmonella and the host cells, and damage of the SCV can lead to bacterial hyper-replication, expulsion of cells from epithelial layers, or induction of apoptotic cell death. Resulting apoptotic bodies may harbour and transmit viable Salmonella to naïve host cells. We recently observed that in contrast to S. enterica serovar Typhimurium (STM), the typhoidal serovar Paratyphi A (SPA) expressed functional flagella after escape from the SCV and is intracellular mobile. We aim to understand the specific intracellular lifestyles of STM and SPA, mechanisms of SCV damage and consequence of cytosolic activities of Salmonella. In particular, we will investigate: • Kinetics of intracellular flagella expression, understanding regulation in SPA vs. STM • Role of Salmonella flagella in SCV damage, induction of host cell responses, and host cell exit • Mechanics of flagella damage to nascent SCV and fate of flagella during invasion We will deploy the live cell reporter EqtSM to monitor SCV damage and to analyse the bacterial and host cell factors contribution to SCV damage or stability: • Application of EqtSM reporter cell lines to analyse SCV damage with high spatial and temporal resolution • Further investigation of Salmonella factors (T3SS effector proteins) involved in SCV damage and stabilization • Set up of improved organoid model systems for analyses of SPA intracellular lifestyle • Generation of optogenetic experimental systems to control flagella functions in infected host cells • Analyses of contribution to vacuolar damage using live cell imaging of infected host cells and minimal system of Salmonella entrapped in giant uni-lamellar vesicles We will investigate how Salmonella can infect host via apoptotic bodies harbouring viable bacteria and address the questions: • Is host cell pyroptotic cell death and release of Salmonella by apoptotic bodies a protected route for infection of naïve host cells? • What is the intracellular environment of STM in AB, and AB after phagocytosis by host cells? The anticipated results will extend our understanding of host pathogen interaction in Salmonella infections, the pathogenesis of typhoidal Salmonella infections, and routes of transmission.