Abstract

Analysis of the genome sequences of the major human bacterial pathogens has provided a large amount of information concerning their metabolic potential. However, our knowledge of the actual metabolic pathways and metabolite fluxes occurring in these pathogens under infection conditions is still limited. In this study, we analysed the intracellular carbon metabolism of enteroinvasive Escherichia coli (EIEC HN280 and EIEC 4608-58) and Salmonella enterica Serovar Typhimurium (Stm 14028) replicating in epithelial colorectal adenocarcinoma cells (Caco-2). To this aim, we supplied [U-13C6]glucose to Caco-2 cells infected with the bacterial strains or mutants thereof impaired in the uptake of glucose, mannose and/or glucose 6-phosphate. The 13C-isotopologue patterns of protein-derived amino acids from the bacteria and the host cells were then determined by mass spectrometry. The data showed that EIEC HN280 growing in the cytosol of the host cells, as well as Stm 14028 replicating in the Salmonella-containing vacuole (SCV) utilised glucose, but not glucose 6-phosphate, other phosphorylated carbohydrates, gluconate or fatty acids as major carbon substrates. EIEC 4608-58 used C3-compound(s) in addition to glucose as carbon source. The labelling patterns reflected strain-dependent carbon flux via glycolysis and/or the Entner-Doudoroff pathway, the pentose phosphate pathway, the TCA cycle and anapleurotic reactions between PEP and oxaloacetate. Mutants of all three strains impaired in the uptake of glucose switched to C3-substrate(s) accompanied by an increased uptake of amino acids (and possibly also other anabolic monomers) from the host cell. Surprisingly, the metabolism of the host cells, as judged by the efficiency of 13C-incorporation into host cell amino acids, was not significantly affected by the infection with either of these intracellular pathogens.

Highlights

  • Enteroinvasive Escherichia coli (EIEC), highly related to Shigella species [1], and Salmonella enterica Serovar Typhimurium

  • The results indicated that in vitro growth of the corresponding mutants of all three strains was strongly impaired in the presence of glucose and glucose-6P, respectively but all mutants were still able to grow in Caco-2 cells albeit with strain-specific reduced efficiencies compared to the wild-type strains

  • We have analysed by 13C-isotopologue profiling the intracellular carbon metabolism of enteroinvasive E. coli (EIEC; closely resembling Shigella species), and S. enterica Serovar Typhimurium

Read more

Summary

Introduction

Enteroinvasive Escherichia coli (EIEC), highly related to Shigella species [1], and Salmonella enterica Serovar Typhimurium Typhimurium in the following) are typical intracellular pathogens, causing bloody diarrhoea and gastroenteritis, respectively, in humans [2] Both microorganisms are mainly food-borne pathogens and reach their target cells after crossing the intestinal epithelium. The genome sequences [5,6] show that these pathogens share a rather similar metabolic setting including all genes for the major central metabolic pathways, i.e. for glycolysis, the Entner-Doudoroff pathway, the pentose phosphate cycle, the citrate (TCA) cycle, the biosyntheses for all amino acids, as well as for the main anapleurotic reactions including the glyoxylate shunt. Transcription of uhpT is very low in the presence of glucose Both pathogens are able to catabolise various C2-, C3-, C4- and C5-substrates and, fatty acids, glycerol, pyruvate, lactate and C4-dicarboxylates may be possible carbon substrates under certain conditions including growth within mammalian host cells

Methods
Results
Conclusion
Full Text
Published version (Free)

Talk to us

Join us for a 30 min session where you can share your feedback and ask us any queries you have

Schedule a call