http://hdl.handle.net/10468/2175 2017-10-30T17:21:02Z http://hdl.handle.net/10468/3559 The aliphatic amidase AmiE is involved in regulation of Pseudomonas aeruginosa virulence Clamens, Thomas; Rosay, Thibaut; Crépin, Alexandre; Grandjean, Teddy; Kentache, Takfarinas; Hardouin, Julie; Bortolotti, Perrine; Neidig, Anke; Mooij, Marlies J.; Hillion, Mélanie; Vieillard, Julien; Cosette, Pascal; Overhage, Joerg; O'Gara, Fergal; Bouffartigues, Emeline; Dufour, Alain; Chevalier, Sylvie; Guery, Benoit; Cornelis, Pierre; Feuilloley, Marc G. J.; Lesouhaitier, Olivier We have previously shown that the eukaryotic C-type natriuretic peptide hormone (CNP) regulates Pseudomonas aeruginosa virulence and biofilm formation after binding on the AmiC sensor, triggering the amiE transcription. Herein, the involvement of the aliphatic amidase AmiE in P. aeruginosa virulence regulation has been investigated. The proteome analysis of an AmiE over-producing strain (AmiE+) revealed an expression change for 138 proteins, including some that are involved in motility, synthesis of quorum sensing compounds and virulence regulation. We observed that the AmiE+ strain produced less biofilm compared to the wild type, and over-produced rhamnolipids. In the same line, AmiE is involved in P. aeruginosa motilities (swarming and twitching) and production of the quorum sensing molecules N-acyl homoserine lactones and Pseudomonas Quinolone Signal (PQS). We observed that AmiE overproduction reduced levels of HCN and pyocyanin causing a decreased virulence in different hosts (i.e. Dictyostelium discoideum and Caenorhabditis elegans). This phenotype was further confirmed in a mouse model of acute lung infection, in which AmiE overproduction resulted in an almost fully virulence decrease. Taken together, our data suggest that, in addition to its role in bacterial secondary metabolism, AmiE is involved in P. aeruginosa virulence regulation by modulating pilus synthesis and cell-to-cell communication. 2017-01-24T00:00:00Z http://hdl.handle.net/10468/3555 Direct and rapid electrochemical detection of Pseudomonas aeruginosa quorum signaling molecules in bacterial cultures and cystic fibrosis sputum samples through cationic surfactant-assisted membrane disruption Buzid, Alyah; Reen, F. Jerry; Langsi, Victor K.; Ó Muimhneacháin, Eoin; O'Gara, Fergal; McGlacken, Gerard P.; Luong, John H. T.; Glennon, Jeremy D. Rapid detection of pathogenic bacteria present in patient samples is of utmost importance for the clinical management of bacterial-induced diseases. Herein, we describe an efficient and direct electrochemical approach for the detection of 2-heptyl-3-hydroxy-4-quinolone (PQS), 2-heptyl-4-hydroxyquinoline (HHQ), and pyocyanin (PYO) as a molecular signature of Pseudomonas aeruginosa (PA), a frequently infecting pathogen with high antibiotic resistance. The cationic surfactant hexadecyltrimethylammonium bromide (CTAB) enhances the effectiveness of an unmodified thin-film boron-doped diamond (BDD) electrode for the direct detection of PYO, HHQ, and PQS in bacterial cultures of PAO1 and PA14. Differential pulse voltammetry (DPV) is then used to monitor the production of these microbial metabolites in bacterial cultures of PAO1 over 10 h without any sample pretreatment. A proposed mechanism for the interaction of CTAB with bacteria cells is examined by zeta (ζ) potential measurements. Furthermore, the detection method is successfully extended to a clinical fluid matrix and applied to PA spiked cystic fibrosis (CF) sputum samples. 2017-01-20T00:00:00Z http://hdl.handle.net/10468/4785 Biotechnological potential of cold adapted Pseudoalteromonas spp. isolated from "deep sea' sponges Borchert, Erik; Knobloch, Stephen; Dwyer, Emilie; Flynn, Sinead; Jackson, Stephen A.; Johannsson, Ragnar; Marteinsson, Viggo T.; O'Gara, Fergal; Dobson, Alan D. W. The marine genus Pseudoalteromonas is known for its versatile biotechnological potential with respect to the production of antimicrobials and enzymes of industrial interest. We have sequenced the genomes of three Pseudoalteromonas sp. strains isolated from different deep sea sponges on the Illumina MiSeq platform. The isolates have been screened for various industrially important enzymes and comparative genomics has been applied to investigate potential relationships between the isolates and their host organisms, while comparing them to free-living Pseudoalteromonas spp. from shallow and deep sea environments. The genomes of the sponge associated Pseudoalteromonas strains contained much lower levels of potential eukaryotic-like proteins which are known to be enriched in symbiotic sponge associated microorganisms, than might be expected for true sponge symbionts. While all the Pseudoalteromonas shared a large distinct subset of genes, nonetheless the number of unique and accessory genes is quite large and defines the pan-genome as open. Enzymatic screens indicate that a vast array of enzyme activities is expressed by the isolates, including -galactosidase, -glucosidase, and protease activities. A -glucosidase gene from one of the Pseudoalteromonas isolates, strain EB27 was heterologously expressed in Escherichia coli and, following biochemical characterization, the recombinant enzyme was found to be cold-adapted, thermolabile, halotolerant, and alkaline active. 2017-01-01T00:00:00Z http://hdl.handle.net/10468/4801 Identification of secondary metabolite gene clusters in the Pseudovibrio genus reveals encouraging biosynthetic potential toward the production of novel bioactive compounds Naughton, Lynn M.; Romano, Stefano; O'Gara, Fergal; Dobson, Alan D. W. Increased incidences of antimicrobial resistance and the emergence of pan-resistant 'superbugs' have provoked an extreme sense of urgency amongst researchers focusing on the discovery of potentially novel antimicrobial compounds. A strategic shift in focus from the terrestrial to the marine environment has resulted in the discovery of a wide variety of structurally and functionally diverse bioactive compounds from numerous marine sources, including sponges. Bacteria found in close association with sponges and other marine invertebrates have recently gained much attention as potential sources of many of these novel bioactive compounds. Members of the genus Pseudovibrio are one such group of organisms. In this study, we interrogate the genomes of 21 Pseudovibrio strains isolated from a variety of marine sources, for the presence, diversity and distribution of biosynthetic gene clusters (BGCs). We expand on results obtained from antiSMASH analysis to demonstrate the similarity between the Pseudovibriorelated BGCs and those characterized in other bacteria and corroborate our findings with phylogenetic analysis. We assess how domain organization of the most abundant type of BGCs present among the isolates (Non-ribosomal peptide synthetases and Polyketide synthases) may influence the diversity of compounds produced by these organisms and highlight for the first time the potential for novel compound production from this genus of bacteria, using a genome guided approach. 2017-01-01T00:00:00Z