Date Published: May 29, 2009
Publisher: Public Library of Science
Author(s): Christopher J. Alteri, Sara N. Smith, Harry L. T. Mobley, Jorge E. Galán.
Microbial pathogenesis studies traditionally encompass dissection of virulence properties such as the bacterium’s ability to elaborate toxins, adhere to and invade host cells, cause tissue damage, or otherwise disrupt normal host immune and cellular functions. In contrast, bacterial metabolism during infection has only been recently appreciated to contribute to persistence as much as their virulence properties. In this study, we used comparative proteomics to investigate the expression of uropathogenic Escherichia coli (UPEC) cytoplasmic proteins during growth in the urinary tract environment and systematic disruption of central metabolic pathways to better understand bacterial metabolism during infection. Using two-dimensional fluorescence difference in gel electrophoresis (2D-DIGE) and tandem mass spectrometry, it was found that UPEC differentially expresses 84 cytoplasmic proteins between growth in LB medium and growth in human urine (P<0.005). Proteins induced during growth in urine included those involved in the import of short peptides and enzymes required for the transport and catabolism of sialic acid, gluconate, and the pentose sugars xylose and arabinose. Proteins required for the biosynthesis of arginine and serine along with the enzyme agmatinase that is used to produce the polyamine putrescine were also up-regulated in urine. To complement these data, we constructed mutants in these genes and created mutants defective in each central metabolic pathway and tested the relative fitness of these UPEC mutants in vivo in an infection model. Import of peptides, gluconeogenesis, and the tricarboxylic acid cycle are required for E. coli fitness during urinary tract infection while glycolysis, both the non-oxidative and oxidative branches of the pentose phosphate pathway, and the Entner-Doudoroff pathway were dispensable in vivo. These findings suggest that peptides and amino acids are the primary carbon source for E. coli during infection of the urinary tract. Because anaplerosis, or using central pathways to replenish metabolic intermediates, is required for UPEC fitness in vivo, we propose that central metabolic pathways of bacteria could be considered critical components of virulence for pathogenic microbes.
Traditional studies of bacterial pathogenesis have focused on pathogen-specific virulence properties including toxins, adhesins, secretion, and iron acquisition systems, and mechanisms to avoid the innate and adaptive immune response. Examining bacterial metabolism during the course of an infection is also critical to further our understanding of pathogenesis and identifying potential targets for new antimicrobial agents. Infectious diseases represent a serious threat to global health because many bacteria that cause disease in humans such as Staphylococcus aureus, Mycobacterium tuberculosis, and E. coli are steadily developing resistance to many of the available treatments –. Since the introduction of antibiotics in the last century, the emergence of bacteria that resist these compounds has rapidly outpaced the discovery and development of new antimicrobial agents . The need to understand bacterial physiology during infection of the host is critical for the development of new antimicrobials or antibiotics that will reduce their burden upon human health.
Bacterial pathogenesis traditionally involves studying virulence traits involved in the production of toxins and effectors, iron acquisition, adherence, invasion, and immune system avoidance. Although many paradigms exist that describe mechanisms of pathogenesis, the contribution of microbial metabolism to bacterial virulence during an infection is less understood. Much work has been done studying E. coli as model organism for characterizing individual central metabolism pathways and enzymes , , –. We have shown here that central metabolism studies in E. coli can be extended to investigate the contribution of central pathways to bacterial pathogenesis using a virulent uropathogenic E. coli strain and a well-established animal model of UTI. It is known that commensal E. coli require the Entner-Doudoroff pathway and glycolysis for colonization in vivo; while the TCA cycle, pentose phosphate pathway, and gluconeogenesis are dispensable in the intestine . In contrast, we have shown that during E. coli infection of the urinary tract, the pathways required for commensal colonization are dispensable while the TCA cycle and gluconeogenesis are necessary for UPEC fitness in vivo. Adaptation to distinct host environments has been previously shown to involve shared traits between commensal and pathogenic strains ,. Because commensal E. coli are an important natural component of the intestine one concern faced when developing antimicrobials that target pathogenic strains is how to avoid eradicating commensal bacteria. Thus, these findings highlight important differences between commensal and pathogenic E. coli that could be exploited for the development of antimicrobials that target these pathways in this pathogen during infections that may not affect commensal strains. Interestingly, in addition to UPEC, gluconeogenesis is required for virulence in microbes that represent an array of pathogenic lifestyles, from intracellular bacteria and parasites ,, plant-pathogenic , and intestinal pathogens ; suggesting that anaplerosis may be a common mechanism of microbial pathogenesis.