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Indole production promotes Escherichia coli mixed-culture growth with Pseudomonas aeruginosa by inhibiting quorum signaling.

ABSTRACT: Indole production by Escherichia coli, discovered in the early 20th century, has been used as a diagnostic marker for distinguishing E. coli from other enteric bacteria. By using transcriptional profiling and competition studies with defined mutants, we show that cyclic AMP (cAMP)-regulated indole formation is a major factor that enables E. coli growth in mixed biofilm and planktonic populations with Pseudomonas aeruginosa. Mutants deficient in cAMP production (cyaA) or the cAMP receptor gene (crp), as well as indole production (tnaA), were not competitive in coculture with P. aeruginosa but could be restored to wild-type competitiveness by supplementation with a physiologically relevant indole concentration. E. coli sdiA mutants, which lacked the receptor for both indole and N-acyl-homoserine lactones (AHLs), showed no change in competitive fitness, suggesting that indole acted directly on P. aeruginosa. An E. coli tnaA mutant strain regained wild-type competiveness if grown with P. aeruginosa AHL synthase (rhlI and rhlI lasI) mutants. In contrast to the wild type, P. aeruginosa AHL synthase mutants were unable to degrade indole. Indole produced during mixed-culture growth inhibited pyocyanin production and other AHL-regulated virulence factors in P. aeruginosa. Mixed-culture growth with P. aeruginosa stimulated indole formation in E. coli cpdA, which is unable to regulate cAMP levels, suggesting the potential for mixed-culture gene activation via cAMP. These findings illustrate how indole, an early described feature of E. coli central metabolism, can play a significant role in mixed-culture survival by inhibiting quorum-regulated competition factors in P. aeruginosa.

SUBMITTER: Chu W

PROVIDER: S-EPMC3255741 | biostudies-literature | 2012 Jan

REPOSITORIES: biostudies-literature

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Indole production promotes Escherichia coli mixed-culture growth with Pseudomonas aeruginosa by inhibiting quorum signaling.

Chu Weihua W Zere Tesfalem R TR Weber Mary M MM Wood Thomas K TK Whiteley Marvin M Hidalgo-Romano Benjamin B Valenzuela Ernesto E McLean Robert J C RJ

Applied and environmental microbiology 20111118 2

Indole production by Escherichia coli, discovered in the early 20th century, has been used as a diagnostic marker for distinguishing E. coli from other enteric bacteria. By using transcriptional profiling and competition studies with defined mutants, we show that cyclic AMP (cAMP)-regulated indole formation is a major factor that enables E. coli growth in mixed biofilm and planktonic populations with Pseudomonas aeruginosa. Mutants deficient in cAMP production (cyaA) or the cAMP receptor gene (c ...[more]

PMID: 22101045

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Project description:Transcriptional profiles of Escherichia coli MG1655 in mixed culture with Pseudomonas aeruginosa PAO1 showed a number of E. coli genes to be upregulated including purA-F and other genes associated with purine synthesis. In contrast, genes associated with pyrimidine synthesis were unaffected. Competition experiments in both planktonic and biofilm cultures, using three purine synthesis mutants, purD, purH, and purT showed little difference in E. coli survival from the parent strain. As purines are components of the cell signals, cAMP and c-di-GMP, we conducted competition experiments with E. coli mutants lacking adenylate cyclase (cyaA), cAMP phosphodiesterase (cpdA), and the catabolite receptor protein (crp), as well as ydeH, an uncharacterized gene that has been associated with c-di-GMP synthesis. Survival of the cyaA and crp mutants during co-culture were significantly less than the parent strain. Supplementation of the media with 1mM cAMP could restore survival of the cyaA mutant but not the crp mutant. In contrast, survival of the cpdA mutant was similar to the parent strain. Survival of the ydeH mutant was moderately less than the parent, suggesting that cAMP has more impact on E. coli mixed culture growth than c-di-GMP. Addition of 1 mM indole restored the survival of both the cyaA and crp mutations. Mutants in genes for tryptophan synthesis (trpE) and indole production (tnaA) showed a loss of competition and recovery through indole supplementation, comparable to the cyaA and crp mutants. Overall, these results suggest indole and cAMP as major contributing factors to E. coli growth in mixed culture. Two replicates of LB grown E. coli ZK126 and P. aeruginosa PAO1 were grown in pure culture at 37C to an OD of 0.3 (log phase) and then mixed 1:1 and incubated for an additional 45 min. RNA was extracted, purified, reverse transcribed to cDNA and then analyzed on E. coli and P. aeruginosa chips from Affymetrix. Expression profiles were compared to pure culture E. coli and P. aeruginosa grown on LB.

Project description:The last decade has witnessed a massive increase in the rate of mortalities caused by multidrug-resistant Pseudomonas aeruginosa. Therefore, developing new strategies to control virulence factors and pathogenicity has received much attention. One of these strategies is quorum sensing inhibition (QSI) which was developed to control Pseudomonas infection. This study aims to validate the effect of one of the most used β-lactam antibiotics; cefoperazone (CFP) and its metallic-derivatives on quorum sensing (QS) and virulence factors of P. aeruginosa. Assessment of quorum sensing inhibitory activity of CFP, cefoperazone Iron complex (CFPF) and cefoperazone Cobalt complex (CFPC) was performed by using reporter strain Chromobacterium violaceum ATCC 12472. Minimal inhibitory concentration (MIC) was carried out by the microbroth dilution method. The influence of sub-MICs (1/4 and 1/2 MICs) of CFP, CFPF and CFPC on virulence factors of P. aeruginosa was evaluated. Data was confirmed on the molecular level by RT-PCR. Also, molecular docking analysis was conducted to figure out the possible mechanisms of QSI. CFP, CFPF, and CFPC inhibited violacein pigment production of C. violaceum ATCC 12472. Sub-MICs of CFP (128- 256 μg/mL), and significantly low concentrations of CFPC (0.5- 16 μg/mL) and CFPF (0.5- 64 μg/mL) reduced the production of QS related virulence factors such as pyocyanin, protease, hemolysin and eliminated biofilm assembly by P. aeruginosa standard strains PAO1 and PA14, and P. aeruginosa clinical isolates Ps1, Ps2, and Ps3, without affecting bacterial viability. In addition, CFP, CFPF, and CFPC significantly reduced the expression of lasI and rhlI genes. The molecular docking analysis elucidated that the QS inhibitory effect was possibly caused by the interaction with QS receptors. Both CFPF and CFPC interacted strongly with LasI, LasR and PqsR receptors with a much high ICM scores compared to CFP that could be the cause of elimination of natural ligand binding. Therefore, CFPC and CFPF are potent inhibitors of quorum sensing signaling and virulence factors of P. aeruginosa.

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Indole production promotes Escherichia coli mixed-culture growth with Pseudomonas aeruginosa by inhibiting quorum signaling.

Publications

Indole production promotes Escherichia coli mixed-culture growth with Pseudomonas aeruginosa by inhibiting quorum signaling.

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