Abstract
BackgroundIn comparison to the comprehensive analyses performed on virulence gene expression, regulation and action, the intracellular metabolism of Salmonella during infection is a relatively under-studied area. We investigated the role of the tricarboxylic acid (TCA) cycle in the intracellular replication of Salmonella Typhimurium in resting and activated macrophages, epithelial cells, and during infection of mice.Methodology/Principal FindingsWe constructed deletion mutations of 5 TCA cycle genes in S. Typhimurium including gltA, mdh, sdhCDAB, sucAB, and sucCD. We found that the mutants exhibited increased net intracellular replication in resting and activated murine macrophages compared to the wild-type. In contrast, an epithelial cell infection model showed that the S. Typhimurium ΔsucCD and ΔgltA strains had reduced net intracellular replication compared to the wild-type. The glyoxylate shunt was not responsible for the net increased replication of the TCA cycle mutants within resting macrophages. We also confirmed that, in a murine infection model, the S. Typhimurium ΔsucAB and ΔsucCD strains are attenuated for virulence.Conclusions/SignificanceOur results suggest that disruption of the TCA cycle increases the ability of S. Typhimurium to survive within resting and activated murine macrophages. In contrast, epithelial cells are non-phagocytic cells and unlike macrophages cannot mount an oxidative and nitrosative defence response against pathogens; our results show that in HeLa cells the S. Typhimurium TCA cycle mutant strains show reduced or no change in intracellular levels compared to the wild-type [1]. The attenuation of the S. Typhimurium ΔsucAB and ΔsucCD mutants in mice, compared to their increased net intracellular replication in resting and activated macrophages suggest that Salmonella may encounter environments within the host where a complete TCA cycle is advantageous.
Highlights
Salmonella enterica is one of the most common food-borne bacterial pathogens and the disease outcomes range from a selflimited gastroenteritis to typhoid fever in mammals
The primary role of the tricarboxylic acid (TCA) cycle is to provide NADH which is used by bacterial cells for ATP synthesis via the electron transport chain (ETC)
Typhimurium Dmdh, DsucAB, DsucCD, and DsdhCDAB strains relative to the wild-type suggested that a complete TCA cycle is not necessary for growth within resting macrophages
Summary
Salmonella enterica is one of the most common food-borne bacterial pathogens and the disease outcomes range from a selflimited gastroenteritis to typhoid fever in mammals. Typhoid infection involves transmission of Salmonella via the ingestion of contaminated food and water followed by bacterial penetration of the small intestinal barrier by invading gut epithelial cells causing bloody diarrhoea. The antimicrobial defences deployed by macrophages include reactive oxygen and reactive nitrosative intermediates (ROI and RNI respectively), as well as antimicrobial peptides [7,8]. The ROI response is bactericidal and occurs approximately 1 h post-infection of macrophages whereas the RNI response is bacteriostatic and occurs approximately 8 h post-infection [9,10,11]. We investigated the role of the tricarboxylic acid (TCA) cycle in the intracellular replication of Salmonella Typhimurium in resting and activated macrophages, epithelial cells, and during infection of mice
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