Dataset Information

Pseudomonas aeruginosa cytotoxicity is attenuated at high cell density and associated with the accumulation of phenylacetic acid.

ABSTRACT:

Background

P. aeruginosa is known to cause acute cytotoxicity against various human and animal cells and tissues.

Methodology/findings

Intriguingly, however, in this study we noticed that while a low cell density inoculum of P. aeruginosa caused severe cytotoxicity against human lung tissue cell line A549, increasing the cell density of bacterial inoculum led to decreased cytotoxicity. Addition of the supernatants from high density bacterial culture to low cell density inoculum protected the human cells from bacterial cytotoxic damage, suggesting that P. aeruginosa may produce and accumulate an inhibitory molecule(s) counteracting its pathogenic infection. The inhibitor was purified from the stationary-phase culture supernatants of P. aeruginosa strain PAO1 using bioassay-guided high performance liquid chromatography (HPLC), and characterized to be phenylacetic acid (PAA) by mass spectrometry and nuclear magnetic resonance spectroscopy. Microarray analysis revealed that treatment of P. aeruginosa with PAA down-regulated the transcriptional expression of Type III secretion system (T3SS) genes and related regulatory genes including rsmA and vfr, which were confirmed by transcriptional and translational analysis.

Conclusions

Identification of bacterial metabolite PAA as a T3SS-specific inhibitor explains this intriguing inverse cell-density-dependent-cytotoxicity phenomenon as T3SS is known to be a key virulence factor associated with cytotoxicity and acute infection. The findings may provide useful clues for design and development of new strategies to combat this formidable bacterial pathogen.

SUBMITTER: Wang J

PROVIDER: S-EPMC3612096 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Publications

Pseudomonas aeruginosa cytotoxicity is attenuated at high cell density and associated with the accumulation of phenylacetic acid.

Wang Jianhe J Dong Yihu Y Zhou Tielin T Liu Xiaoling X Deng Yinyue Y Wang Chao C Lee Jasmine J Zhang Lian-Hui LH

PloS one 20130329 3

<h4>Background</h4>P. aeruginosa is known to cause acute cytotoxicity against various human and animal cells and tissues.<h4>Methodology/findings</h4>Intriguingly, however, in this study we noticed that while a low cell density inoculum of P. aeruginosa caused severe cytotoxicity against human lung tissue cell line A549, increasing the cell density of bacterial inoculum led to decreased cytotoxicity. Addition of the supernatants from high density bacterial culture to low cell density inoculum pr ...[more]

PMID: 23555919

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Project description:Styrene is a toxic and potentially carcinogenic alkenylbenzene used extensively in the polymer processing industry. Significant quantities of contaminated liquid waste are generated annually as a consequence. However, styrene is not a true xenobiotic and microbial pathways for its aerobic assimilation, via an intermediate, phenylacetic acid, have been identified in a diverse range of environmental isolates. The potential for microbial bioremediation of styrene waste has received considerable research attention over the last number of years. As a result the structure, organisation and encoded function of the genes responsible for styrene and phenylacetic acid sensing, uptake and catabolism have been elucidated. However, a limited understanding persists in relation to host specific regulatory molecules which may impart additional control over these pathways. In this study the styrene degrader Pseudomonas putida CA-3 was subjected to random mini-Tn5 mutagenesis and mutants screened for altered styrene/phenylacetic acid utilisation profiles potentially linked to non-catabolon encoded regulatory influences.One mutant, D7, capable of growth on styrene, but not on phenylacetic acid, harboured a Tn5 insertion in the rpoN gene encoding ?54. Complementation of the D7 mutant with the wild type rpoN gene restored the ability of this strain to utilise phenylacetic acid as a sole carbon source. Subsequent RT-PCR analyses revealed that a phenylacetate permease, PaaL, was expressed in wild type P. putida CA-3 cells utilising styrene or phenylacetic acid, but could not be detected in the disrupted D7 mutant. Expression of plasmid borne paaL in mutant D7 was found to fully restore the phenylacetic acid utilisation capacity of the strain to wild type levels. Bioinformatic analysis of the paaL promoter from P. putida CA-3 revealed two ?54 consensus binding sites in a non-archetypal configuration, with the transcriptional start site being resolved by primer extension analysis. Comparative analyses of genomes encoding phenylacetyl CoA, (PACoA), catabolic operons identified a common association among styrene degradation linked PACoA catabolons in Pseudomonas species studied to date.In summary, this is the first study to report RpoN dependent transcriptional activation of the PACoA catabolon paaL gene, encoding a transport protein essential for phenylacetic acid utilisation in P. putida CA-3. Bioinformatic analysis is provided to suggest this regulatory link may be common among styrene degrading Pseudomonads.

Dataset Information

Pseudomonas aeruginosa cytotoxicity is attenuated at high cell density and associated with the accumulation of phenylacetic acid.

Background

Methodology/findings

Conclusions

Publications

Pseudomonas aeruginosa cytotoxicity is attenuated at high cell density and associated with the accumulation of phenylacetic acid.

Similar Datasets

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