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Engineering microbes to sense and eradicate Pseudomonas aeruginosa, a human pathogen.

ABSTRACT: Synthetic biology aims to systematically design and construct novel biological systems that address energy, environment, and health issues. Herein, we describe the development of a synthetic genetic system, which comprises quorum sensing, killing, and lysing devices, that enables Escherichia coli to sense and kill a pathogenic Pseudomonas aeruginosa strain through the production and release of pyocin. The sensing, killing, and lysing devices were characterized to elucidate their detection, antimicrobial and pyocin release functionalities, which subsequently aided in the construction of the final system and the verification of its designed behavior. We demonstrated that our engineered E. coli sensed and killed planktonic P. aeruginosa, evidenced by 99% reduction in the viable cells. Moreover, we showed that our engineered E. coli inhibited the formation of P. aeruginosa biofilm by close to 90%, leading to much sparser and thinner biofilm matrices. These results suggest that E. coli carrying our synthetic genetic system may provide a novel synthetic biology-driven antimicrobial strategy that could potentially be applied to fighting P. aeruginosa and other infectious pathogens.

SUBMITTER: Saeidi N

PROVIDER: S-EPMC3202794 | biostudies-literature | 2011 Aug

REPOSITORIES: biostudies-literature

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Engineering microbes to sense and eradicate Pseudomonas aeruginosa, a human pathogen.

Saeidi Nazanin N Wong Choon Kit CK Lo Tat-Ming TM Nguyen Hung Xuan HX Ling Hua H Leong Susanna Su Jan SS Poh Chueh Loo CL Chang Matthew Wook MW

Molecular systems biology 20110816

Synthetic biology aims to systematically design and construct novel biological systems that address energy, environment, and health issues. Herein, we describe the development of a synthetic genetic system, which comprises quorum sensing, killing, and lysing devices, that enables Escherichia coli to sense and kill a pathogenic Pseudomonas aeruginosa strain through the production and release of pyocin. The sensing, killing, and lysing devices were characterized to elucidate their detection, antim ...[more]

PMID: 21847113

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Project description:UnlabelledPseudomonas aeruginosa uses N-acyl-homoserine lactone (AHL)-dependent quorum sensing (QS) systems to control the expression of secreted effectors. These effectors can be crucial to the ecological fitness of the bacterium, playing roles in nutrient acquisition, microbial competition, and virulence. In this study, we investigated the metabolic consequences of AHL-dependent QS by monitoring the metabolic profile(s) of a lasI rhlI double mutant (unable to make QS signaling molecules) and its wild-type progenitor as they progressed through the growth curve. Analysis of culture supernatants by (1)H-nuclear magnetic resonance ((1)H-NMR) spectroscopy revealed that at the point where AHL concentrations peaked in the wild type, the metabolic footprints (i.e., extracellular metabolites) of the wild-type and lasI rhlI mutant diverged. Subsequent gas chromatography-mass spectrometry (GC-MS)-based analysis of the intracellular metabolome revealed QS-dependent perturbations in around one-third of all identified metabolites, including altered concentrations of tricarboxylic acid (TCA) cycle intermediates, amino acids, and fatty acids. Further targeted fatty acid methyl ester (FAME) GC-MS-based profiling of the cellular total fatty acid pools revealed that QS leads to changes associated with decreased membrane fluidity and higher chemical stability. However, not all of the changes we observed were necessarily a direct consequence of QS; liquid chromatography (LC)-MS analyses revealed that polyamine levels were elevated in the lasI rhlI mutant, perhaps a response to the absence of QS-dependent adaptations. Our data suggest that QS leads to a global readjustment in central metabolism and provide new insight into the metabolic changes associated with QS during stationary-phase adaptation.ImportanceQuorum sensing (QS) is a transcriptional regulatory mechanism that allows bacteria to coordinate their gene expression profile with the population cell density. The opportunistic human pathogen Pseudomonas aeruginosa uses QS to control the production of secreted virulence factors. In this study, we show that QS elicits a global "metabolic rewiring" in P. aeruginosa. This metabolic rerouting of fluxes is consistent with a variety of drivers, ranging from altered QS-dependent transcription of "metabolic genes" through to the effect(s) of global "metabolic readjustment" as a consequence of QS-dependent exoproduct synthesis, as well as a general stress response, among others. To our knowledge, this is the first study of its kind to assess the global impact of QS on the metabolome.

Project description:Histamine is a key biological signaling molecule. It acts as a neurotransmitter in the central and peripheral nervous systems and coordinates local inflammatory responses by modulating the activity of different immune cells. During inflammatory processes, including bacterial infections, neutrophils stimulate the production and release of histamine. Here, we report that the opportunistic human pathogen Pseudomonas aeruginosa exhibits chemotaxis toward histamine. This chemotactic response is mediated by the concerted action of the TlpQ, PctA, and PctC chemoreceptors, which display differing sensitivities to histamine. Low concentrations of histamine were sufficient to activate TlpQ, which binds histamine with an affinity of 639?nM. To explore this binding, we resolved the high-resolution structure of the TlpQ ligand binding domain in complex with histamine. It has an unusually large dCACHE domain and binds histamine through a highly negatively charged pocket at its membrane distal module. Chemotaxis to histamine may play a role in the virulence of P. aeruginosa by recruiting cells at the infection site and consequently modulating the expression of quorum-sensing-dependent virulence genes. TlpQ is the first bacterial histamine receptor to be described and greatly differs from human histamine receptors, indicating that eukaryotes and bacteria have pursued different strategies for histamine recognition.IMPORTANCE Genome analyses indicate that many bacteria possess an elevated number of chemoreceptors, suggesting that these species are able to perform chemotaxis to a wide variety of compounds. The scientific community is now only beginning to explore this diversity and to elucidate the corresponding physiological relevance. The discovery of histamine chemotaxis in the human pathogen Pseudomonas aeruginosa provides insight into tactic movements that occur within the host. Since histamine is released in response to bacterial pathogens, histamine chemotaxis may permit bacterial migration and accumulation at infection sites, potentially modulating, in turn, quorum-sensing-mediated processes and the expression of virulence genes. As a consequence, the modulation of histamine chemotaxis by signal analogues may result in alterations of the bacterial virulence. As the first report of bacterial histamine chemotaxis, this study lays the foundation for the exploration of the physiological relevance of histamine chemotaxis and its role in pathogenicity.

Dataset Information

Engineering microbes to sense and eradicate Pseudomonas aeruginosa, a human pathogen.

Publications

Engineering microbes to sense and eradicate Pseudomonas aeruginosa, a human pathogen.

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