Role of Host Cell-Derived Amino Acids in Nutrition of Intracellular Salmonella enterica

Abstract

The facultative intracellular pathogen Salmonella enterica resides in a specific membrane-bound compartment termed the Salmonella-containing vacuole (SCV). Despite being segregated from access to metabolites in the host cell cytosol, Salmonella is able to efficiently proliferate within the SCV. We set out to unravel the nutritional supply of Salmonella in the SCV with focus on amino acids. We studied the availability of amino acids by the generation of auxotrophic strains for alanine, asparagine, aspartate, glutamine, and proline in a macrophage cell line (RAW264.7) and an epithelial cell line (HeLa) and examined access to extracellular nutrients for nutrition. Auxotrophies for alanine, asparagine, or proline attenuated intracellular replication in HeLa cells, while aspartate, asparagine, or proline auxotrophies attenuated intracellular replication in RAW264.7 macrophages. The different patterns of intracellular attenuation of alanine-or aspartate-auxotrophic strains support distinct nutritional conditions in HeLa cells and RAW264.7 macrophages. Supplementation of medium with individual amino acids restored the intracellular replication of mutant strains auxotrophic for asparagine, proline, or glutamine. Similarly, a mutant strain deficient in succinate dehydrogenase was complemented by the extracellular addition of succinate. Complementation of the intracellular replication of auxotrophic Salmonella by external amino acids was possible if bacteria were proficient in the induction of Salmonella-induced filaments (SIFs) but failed in a SIF-deficient background. We propose that the ability of intracellular Salmonella to redirect host cell vesicular transport provides access of amino acids to auxotrophic strains and, more generally, is essential to continuously supply bacteria within the SCV with nutrients.T he facultative intracellular life-style is a common virulence strategy among bacterial pathogens, and intracellular lifestyles are as diverse as the diseases caused by these pathogens. While some bacteria lyse the host cell membrane compartment and initiate replication in the cytosol, others remain in a host cell-derived membrane compartment that is modified to allow intravacuolar survival and replication. To understand the lifestyle of bacterial pathogens, it is of central importance to analyze which nutritional limitations are experienced by the pathogen in its intracellular habitat and how the pathogen adapts its metabolism in order to survive and proliferate despite these limitations.Salmonella enterica is an invasive, facultative, intracellular pathogen responsible for foodborne diseases ranging from localized gastroenteritis to systemic typhoid fever. Inside mammalian host cells, S. enterica resides in a specialized membrane-bound compartment, the Salmonella-containing vacuole (SCV). The SCV is modified by the action of virulence factors of S. enterica, and the function of the type III secretion system (T3SS) encoded by Salmonella pathogenicity island 2 (SPI2) and its effector proteins is of central impor...

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