SifA-mediated Remodeling of the Salmonella -Containing Vacuole Prevents Bacterial Dormancy by Promoting Nutrient Accessibility

Salmonella exploits a range of intracellular niches within host cells, adapting to different microenvironments that support its survival and replication. This niche diversity is mediated by bacterial effectors injected via the two type 3 secretion systems (T3SS). Salmonella resides either within membrane-bound Salmonella containing vacuoles (SCV) or they grow rapidly within the cytosol upon SCV rupture. Recently, we identified a third intracellular subpopulation, dormant Salmonella within modified vacuoles in epithelial cells, which can survive for up to 7 days in vitro . To explore how bacterial effectors influence the balance of these three subpopulations, we constructed a panel of mutants lacking genes encoding key effectors and examined their intracellular behaviors at 6 and 24 hours post-infection (hpi). Deletion of the T3SS-2 effector SifA significantly increased the dormant subpopulation at later infection time points, identifying SifA as the first known effector regulating bacterial dormancy in epithelial cells. SifA-induced Salmonella -induced filaments (SIFs) characterize the mature Salmonella -containing vacuole (SCV), and we observed that SIFs were needed for intravacuolar growth of Salmonella . We could show that SIF formation was critical for nutrient acquisition within the SCV; inhibition of SIF formation resulted in a higher proportion of dormant bacteria, akin to the effect of reduced glucose availability during infection. Both bacterial and host glycolysis pathways were required to prevent dormancy, as proper nutrient scavenging through SIF-mediated modification of the SCV is essential for maintaining a replicative, non-dormant state. These results underscore the importance of nutrient access, facilitated by reprogramming host endomembrane trafficking, for Salmonella to avoid dormancy and sustain active replication within its intracellular niche.