Listeria monocytogenes cell-to-cell spread bypasses nutrient limitation for replicating intracellular bacteria

Listeria monocytogenes is an intracellular bacterial pathogen that obtains nutrients from the mammalian host cell to fuel its replication in cytosol. Sparse infection of epithelial monolayers by L. monocytogenes results in the formation of distinct infectious foci, where each focus originates from the initial infection of a single host cell followed by multiple rounds of active bacterial cell-to-cell spread into neighboring host cells in the monolayer. We used time-lapse microscopy to measure changes in bacterial growth rate in individual foci over time and found that intracellular bacteria initially replicate exponentially, but then bacterial growth rate slows later in infection, particularly in the center of the infectious focus. We found that the intracellular replication rate of L. monocytogenes is measurably decreased by limiting host cell glucose availability, by decreasing the rate of intracellular bacterial oligopeptide import, and, most interestingly, by alterations in host cell junctional proteins that limit bacterial spread into neighboring cells without directly affecting bacterial growth or metabolism. By measuring the carrying capacity of individual host cells, we found that the nutritional density of cytoplasm is comparable to rich medium. Taken together, our results indicate that the rate of intracellular L. monocytogenes replication is governed by a balance of the rate of nutrient depletion by the bacteria, the rate of nutrient replenishment by the metabolically active host cells, and the rate of bacterial cell-to-cell spread which enables the bacteria to seek out "greener pastures" before nutrient availability in a single host cell becomes limiting. SIGNIFICANCEListeria monocytogenes is a foodborne pathogen that is capable of facultative intracellular growth in a wide variety of mammalian cell types. It uses actin-based motility to spread directly from the cytoplasm of one infected host cell into another. After initially infecting cells in the small intestine, L. monocytogenes can circulate inside of infected macrophages, and subsequently spread to cells of distal organs, where it can cause spontaneous abortions in pregnant women and meningitis in immunocompromised individuals. In addition to facilitating body-wide dissemination, the actin-based cell-to-cell spread of L. monocytogenes contributes to the ability of the pathogen to evade the cell-mediated arm of the immune system. In this work, we find that cell-to-cell spread also benefits the bacteria in a third way, by enabling the bacteria to overcome the nutrient limitation they would face if they remained confined to a single host cell, such that they never enter stationary phase.