Anaerobic Survival of Pseudomonas aeruginosa by Pyruvate Fermentation Requires an Usp-Type Stress Protein

ABSTRACT Recently, we identified a pyruvate fermentation pathway in Pseudomonas aeruginosa sustaining anaerobic survival in the absence of alternative anaerobic respiratory and fermentative energy generation systems (M. Eschbach, K. Schreiber, K. Trunk, J. Buer, D. Jahn, and M. Schobert, J. Bacteriol. 186:4596-4604, 2004). Anaerobic long-term survival of P. aeruginosa might be essential for survival in deeper layers of a biofilm and the persistent infection of anaerobic mucus plaques in the cystic fibrosis lung. Proteome analysis of P. aeruginosa cells during a 7-day period of pyruvate fermentation revealed the induced synthesis of three enzymes involved in arginine fermentation, ArcA, ArcB, and ArcC, and the outer membrane protein OprL. Moreover, formation of two proteins of unknown function, PA3309 and PA4352, increased by factors of 72- and 22-fold, respectively. Both belong to the group of universal stress proteins (Usp). Long-term survival of a PA3309 knockout mutant by pyruvate fermentation was found drastically reduced. The oxygen-sensing regulator Anr controls expression of the P PA3309-lacZ reporter gene fusion after a shift to anaerobic conditions and further pyruvate fermentation. PA3309 expression was also found induced during the anaerobic and aerobic stationary phases. This aerobic stationary-phase induction is independent of the regulatory proteins Anr, RpoS, RelA, GacA, RhlR, and LasR, indicating a currently unknown mechanism of stationary-phase-dependent gene activation. PA3309 promoter activity was detected in the deeper layers of a P. aeruginosa biofilm using a P PA3309-gfp (green fluorescent protein gene) fusion and confocal laser-scanning microscopy. This is the first description of an Anr-dependent, anaerobically induced, and functional Usp-like protein in bacteria.

Download Full-text

Long-Term Anaerobic Survival of the Opportunistic Pathogen Pseudomonas aeruginosa via Pyruvate Fermentation

Journal of Bacteriology ◽

10.1128/jb.186.14.4596-4604.2004 ◽

2004 ◽

Vol 186 (14) ◽

pp. 4596-4604 ◽

Cited By ~ 149

Author(s):

Martin Eschbach ◽

Kerstin Schreiber ◽

Katharina Trunk ◽

Jan Buer ◽

Dieter Jahn ◽

...

Keyword(s):

Pseudomonas Aeruginosa ◽

Opportunistic Pathogen ◽

Integration Host Factor ◽

Anaerobic Growth ◽

Nitrate Respiration ◽

Term Survival ◽

Long Term Survival ◽

Pyruvate Fermentation ◽

Activity Measurements

ABSTRACT Denitrification and arginine fermentation are central metabolic processes performed by the opportunistic pathogen Pseudomonas aeruginosa during biofilm formation and infection of lungs of patients with cystic fibrosis. Genome-wide searches for additional components of the anaerobic metabolism identified potential genes for pyruvate-metabolizing NADH-dependent lactate dehydrogenase (ldhA), phosphotransacetylase (pta), and acetate kinase (ackA). While pyruvate fermentation alone does not sustain significant anaerobic growth of P. aeruginosa, it provides the bacterium with the metabolic capacity for long-term survival of up to 18 days. Detected conversion of pyruvate to lactate and acetate is dependent on the presence of intact ldhA and ackA-pta loci, respectively. DNA microarray studies in combination with reporter gene fusion analysis and enzyme activity measurements demonstrated the anr- and ihfA-dependent anaerobic induction of the ackA-pta promoter. Potential Anr and integration host factor binding sites were localized. Pyruvate-dependent anaerobic long-term survival was found to be significantly reduced in anr and ihfA mutants. No obvious ldhA regulation by oxygen tension was observed. Pyruvate fermentation is pH dependent. Nitrate respiration abolished pyruvate fermentation, while arginine fermentation occurs independently of pyruvate utilization.

Download Full-text

Identification of genes involved in long-term survival in Pseudomonas aeruginosa PAO1

African Journal of Microbiology Research ◽

10.5897/ajmr12.2283 ◽

2012 ◽

Vol 6 (50) ◽

pp. 7579-7586

Author(s):

Kong Weina ◽

Chen Lin ◽

Wang Yue

Keyword(s):

Pseudomonas Aeruginosa ◽

Term Survival ◽

Long Term Survival ◽

Pseudomonas Aeruginosa Pao1

Download Full-text

A metabolic strategy to enhance long-term survival by Phx1 through stationary phase-specific pyruvate decarboxylases in fission yeast

Aging ◽

10.18632/aging.100682 ◽

2014 ◽

Vol 6 (7) ◽

pp. 587-601 ◽

Cited By ~ 4

Author(s):

Ji-Yoon Kim ◽

Eun-Jung Kim ◽

Luis Lopez-Maury ◽

Jürg Bähler ◽

Jung-Hye Roe

Keyword(s):

Stationary Phase ◽

Fission Yeast ◽

Term Survival ◽

Long Term Survival

Download Full-text

UDP-glucose pyrophosphorylase Ugp1 is involved in oxidative stress response and long-term survival during stationary phase in Saccharomyces cerevisiae

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2015.10.090 ◽

2015 ◽

Vol 467 (4) ◽

pp. 657-663 ◽

Cited By ~ 7

Author(s):

Dae-Gwan Yi ◽

Won-Ki Huh

Keyword(s):

Oxidative Stress ◽

Saccharomyces Cerevisiae ◽

Stress Response ◽

Stationary Phase ◽

Oxidative Stress Response ◽

Term Survival ◽

Long Term Survival

Download Full-text

Culture Volume and Vessel Affect Long-Term Survival, Mutation Frequency, and Oxidative Stress of Escherichia coli

Applied and Environmental Microbiology ◽

10.1128/aem.03150-13 ◽

2013 ◽

Vol 80 (5) ◽

pp. 1732-1738 ◽

Cited By ~ 32

Author(s):

Karin E. Kram ◽

Steven E. Finkel

Keyword(s):

Oxidative Stress ◽

Escherichia Coli ◽

Stationary Phase ◽

Mutation Frequency ◽

Physiological Responses ◽

Culture Vessel ◽

Term Survival ◽

Long Term Survival ◽

Content Type

ABSTRACTBacteria such asEscherichia coliare frequently studied during exponential- and stationary-phase growth. However, many strains can survive in long-term stationary phase (LTSP), without the addition of nutrients, from days to several years. During LTSP, cells experience a variety of stressors, including reactive oxidative species, nutrient depletion, and metabolic toxin buildup, that lead to physiological responses and changes in genetic stability. In this study, we monitored survival during LTSP, as well as reporters of genetic and physiological change, to determine how the physical environment affectsE. coliduring long-term batch culture. We demonstrate differences in yield during LTSP in cells incubated in LB medium in test tubes versus Erlenmeyer flasks, as well as growth in different volumes of medium. We determined that these differences are only partially due to differences in oxygen levels by incubating the cells in different volumes of media under anaerobic conditions. Since we hypothesized that differences in long-term survival are the result of changes in physiological outputs during the late log and early stationary phases, we monitored alkalization, mutation frequency, oxidative stress response, and glycation. Although initial cell yields are essentially equivalent under each condition tested, physiological responses vary greatly in response to culture environment. Incubation in lower-volume cultures leads to higheroxyRexpression but lower mutation frequency and glycation levels, whereas incubation in high-volume cultures has the opposite effect. We show here that even under commonly used experimental conditions that are frequently treated as equivalent, the stresses experienced by cells can differ greatly, suggesting that culture vessel and incubation conditions should be carefully considered in the planning or analysis of experiments.

Download Full-text

Long-term survival during stationary phase: evolution and the GASP phenotype

Nature Reviews Microbiology ◽

10.1038/nrmicro1340 ◽

2006 ◽

Vol 4 (2) ◽

pp. 113-120 ◽

Cited By ~ 312

Author(s):

Steven E. Finkel

Keyword(s):

Stationary Phase ◽

Phase Evolution ◽

Term Survival ◽

Long Term Survival

Download Full-text

Recovery from long-term stationary phase and stress survival in Escherichia coli require the l-isoaspartyl protein carboxyl methyltransferase at alkaline pH

Microbiology ◽

10.1099/mic.0.27835-0 ◽

2005 ◽

Vol 151 (7) ◽

pp. 2151-2158 ◽

Cited By ~ 17

Author(s):

Wade M. Hicks ◽

Matthew V. Kotlajich ◽

Jonathan E. Visick

Keyword(s):

Escherichia Coli ◽

Stationary Phase ◽

Alkaline Ph ◽

Term Survival ◽

Long Term Survival ◽

E Coli ◽

Carboxyl Methyltransferase ◽

Protein Carboxyl Methyltransferase ◽

Stress Survival

The l-isoaspartyl protein carboxyl methyltransferase (pcm) can stimulate repair of isoaspartyl residues arising spontaneously in proteins to normal l-aspartyl residues. PCM is needed in Escherichia coli for maximal long-term survival when exposed to oxidative stress, osmotic stress, repeated heat stress or methanol. The effect of pH on a pcm mutant during long-term stationary phase was examined. PCM was not required for long-term survival of E. coli subjected to pH stress alone; however, PCM-deficient cells showed impaired resistance to paraquat and methanol only at elevated pH. The mutant also showed stress-survival phenotypes in minimal medium buffered to pH 9·0. Accumulation of isoaspartyl residues was accelerated at pH 8·0 or 9·0 in vivo, though PCM-deficient cells did not show higher levels of damage. However, the pcm mutant displayed an extended lag phase in recovering from stationary phase at pH 9·0. Protein repair by PCM thus plays a key role in long-term stress survival only at alkaline pH in E. coli, and it may function primarily to repair damage in cells that are recovering from nutrient limitation and in those cells that are able to divide during long-term stationary phase.

Download Full-text

Dual transcriptional analysis reveals adaptation of host and pathogen to intracellular survival of Pseudomonas aeruginosa associated with urinary tract infection

PLoS Pathogens ◽

10.1371/journal.ppat.1009534 ◽

2021 ◽

Vol 17 (4) ◽

pp. e1009534

Author(s):

Cristina Penaranda ◽

Nicole M. Chumbler ◽

Deborah T. Hung

Keyword(s):

Pseudomonas Aeruginosa ◽

Urinary Tract Infection ◽

Urinary Tract ◽

Tract Infection ◽

Transcriptional Response ◽

Intracellular Survival ◽

Host Cells ◽

Term Survival ◽

Long Term Survival

Long-term survival of bacterial pathogens during persistent bacterial infections can be associated with antibiotic treatment failure and poses a serious public health problem. Infections caused by the Gram-negative pathogen Pseudomonas aeruginosa, which can cause both acute and chronic infections, are particularly challenging due to its high intrinsic resistance to antibiotics. The ineffectiveness of antibiotics is exacerbated when bacteria reside intracellularly within host cells where they can adopt a drug tolerant state. While the early steps of adherence and entry of P. aeruginosa into mammalian cells have been described, the subsequent fate of internalized bacteria, as well as host and bacterial molecular pathways facilitating bacterial long-term survival, are not well defined. In particular, long-term survival within bladder epithelial cells has not been demonstrated and this may have important implications for the understanding and treatment of UTIs caused by P. aeruginosa. Here, we demonstrate and characterize the intracellular survival of wild type (WT) P. aeruginosa inside bladder epithelial cells and a mutant with a disruption in the bacterial two-component regulator AlgR that is unable to survive intracellularly. Using simultaneous dual RNA-seq transcriptional profiling, we define the transcriptional response of intracellular bacteria and their corresponding invaded host cells. The bacterial transcriptional response demonstrates that WT bacteria rapidly adapt to the stress encountered in the intracellular environment in contrast to ΔalgR bacteria. Analysis of the host transcriptional response to invasion suggests that the NF-ΚB signaling pathway, previously shown to be required for extracellular bacterial clearance, is paradoxically also required for intracellular bacterial survival. Lastly, we demonstrate that intracellular survival is important for pathogenesis of P. aeruginosa in vivo using a model of murine urinary tract infection. We propose that the unappreciated ability of P. aeruginosa to survive intracellularly may play an important role in contributing to the chronicity and recurrence of P. aeruginosa in urinary tract infections.

Download Full-text