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1
Turner, Keith Holte. "Bistability in Pseudomonas aeruginosa." Thesis, Harvard University, 2012. http://dissertations.umi.com/gsas.harvard:10159.
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The opportunistic pathogen P. aeruginosa is a leading cause of hospital-accquired infections, and is also the primary cause of morbidity and mortality in patients with cystic fibrosis (CF). In this thesis, I describe the identification and characterization of a novel LysR-type transcription regulator (LTTR) of P. aeruginosa named BexR. I show that BexR exhibits reversible ON/OFF bistable expression, which leads to the bistable expression of several genes including one encoding a virulence factor. I present results suggesting that this bistable expression depends on positive feedback of BexR. This work illuminates the simplicity with which a transcription regulatory network can exhibit a complex behavior and generate phenotypic diversity in a clonal population.
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Silistre, Hazel. "Riboregulation in Pseudomonas aeruginosa." Thesis, University of Nottingham, 2016. http://eprints.nottingham.ac.uk/32634/.
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The opportunistic human pathogen Pseudomonas aeruginosa controls virulence, production of secondary metabolites, motility, biofilm formation, growth in anaerobic conditions, intracellular and intercellular signalling and the switch from an acute to a chronic mode of infection at the transcriptional and post-transcriptional levels by modulation of the Gac/Rsm system. Cell density-dependent signal accumulation and environmental stimulators such as pH changes and ion limitation activate the GacS/GacA two-component system which in turn triggers transcription of the small regulatory RNAs RsmY and RsmZ. These sRNAs sequester multiple copies of the RNA-binding protein RsmA, antagonising its function. The RsmA/CsrA proteins act as translational repressors by binding to the GGA-motifs in the untranslated region of target mRNAs and blocking ribosome binding. In this study, the biological function of RsmN, an RsmA homologue with a conserved RNA-binding pocket but a distinct protein folding, the predicted autoregulatory mechanism of RsmN, the nature of target transcripts of RsmN, and the cross-regulation between the two Rsm proteins were investigated. The positive control of proteolytic and elastinolytic activities and swarming motility by RsmN has been demonstrated using single and inducible double deletion mutants of rsmN. Furthermore, rsmN deletion increased microcolony formation during biofilm formation. Regulation by RsmN was most apparent in the absence of RsmA, when rsmN expression was induced via a multicopy plasmid and at temperatures lower than 37°C. The double deletion of rsmA and rsmN affected growth, diminished proteolytic and elastinolytic activities, triggered autolysis and led to the increased secretion of the type VI secretion system protein Hcp1. Moreover, the double deletion of rsmA and rsmN altered the colony morphology of P. aeruginosa. Mutagenesis of the functionally critical, conserved RNA-binding residue which is identified as Arg44 in RsmA and Arg62 in RsmN resulted in the loss of RsmN function. In a genome-wide analysis by RNASeq, target transcripts were co-purified with RsmN from 37°C and 34°C cultures of a wild-type strain expressing rsmN in multicopy numbers. RNASeq results indicated that small regulatory RNAs such as CrcZ, RsmY and RgsA are common targets of RsmN and RsmA, whereas PhrS is a target of RsmN only. Other common RsmA and RsmN targets included transcriptional regulators, heat shock proteins, proteases, starvation response proteins, components of the denitrification pathway, outer membrane proteins required for pore formation, type III and type VI secretion system proteins and RsmA. Transcripts of heat shock proteins, the tss operon genes and rsmA were enriched by RsmN at 37°C but not at 34°C whereas the lasB transcript was enriched by RsmN at 34°C but not at 37°C. Based on the list of common targets of RsmA and RsmN and the results obtained from phenotypic assays, induction of the lytic Pf4 prophage, accumulation of alkyl quinolone or c-di-GMP signalling molecules, imbalanced redox state, carbon starvation, increased membrane permeability, and aggregation of misfolded proteins are suggested as possible mechanisms triggering the excessive autolysis of the rsmNind ΔrsmA mutant under uninducing conditions. The data gathered so far suggests that rsmN is differentially expressed, with increased RsmN activity at temperatures below 37°C in comparison with RsmA, and, RsmA and RsmN collectively contribute to the regulation of secondary metabolite production, motility and microcolony formation in P. aeruginosa.
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Daly, Philip J. "Permeability of pseudomonas aeruginosa." Thesis, Aston University, 1986. http://publications.aston.ac.uk/12458/.
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Eschbach, Martin. "Molekulare Regulation und Biochemie des anaeroben Langzeitüberlebens von Pseudomonas aeruginosa." [S.l. : s.n.], 2004. http://deposit.ddb.de/cgi-bin/dokserv?idn=971750645.
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Kluftinger, Janet Louise. "Macrophage interaction with Pseudomonas aeruginosa." Thesis, University of British Columbia, 1988. http://hdl.handle.net/2429/29129.
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The interactions of macrophages with Pseudomonas aeruginosa were studied. Five monoclonal antibodies specific for porin protein F were tested for their ability to opsonize P. aeruginosa for complement-independent phagocytosis by unelicited mouse peritoneal macrophages, human peripheral blood monocytes and mouse macrophage cell line P388[sub D1]. All five antibodies significantly increased the level of bacterial uptake over that obtained with the non-opsonic controls. The relative effectiveness of the different antibodies was approximately the same in all cell types indicating that the P388[sub D1] cells can be used as a model for normal macrophages. Of the four monoclonal antibodies directed against similar epitopes of protein F, the three IgGl monoclonal antibodies were substantially more opsonic than the one IgG2a isotype.
P. aeruginosa cytotoxin and periplasmic contents caused a significant reduction in antibody-mediated phagocytosis of P. aeruginosa. Phagocytosis was restored upon pre-incubation with anti-cytotoxin serum. Both cytotoxin and periplasmic contents caused depolarization of the P388[sub D1] cell membrane, as demonstrated using a polarization-sensitive fluorescent probe. These data indicated that P. aeruginosa cytotoxin was localized in the periplasm and had the potential to inhibit macrophage-mediated phagocytosis, possibly by perturbing ion gradients across the macrophage plasma membrane. Monoclonal antibodies directed against protein F were also capable of enhancing phagocytosis of in vivo-grown P. aeruginosa. P. aeruginosa cells taken directly from the in vivo growth system were significantly more susceptible to macrophage phagocytosis than were the same cells after being washed in buffer. The phagocytosis-promoting factor could be isolated from the supernatant of in vivo-grown bacteria and was determined to be fibronectin. Data indicated that promotion by fibronectin of non-opsonic phagocytosis was mediated by direct activation of the macrophages. The tetrapeptide arginine-glycine-aspartate-serine in the eukaryotic cell binding domain of fibronectin was demonstrated to be the macrophage-activating region. Phagocytosis of a mutant P. aeruginosa strain lacking surface pili could not be enhanced by fibronectin. Furthermore, exogenously added Pseudomonas pili was capable of abrogating the enhanced phagocytosis of the wild type strain observed with fibronectin-activated macrophages. It was concluded that Pseudomonas pili were the bacterial ligands required for attachment to fibronectin-activated macrophages in the initial stages of non-opsonic phagocytosis.Science, Faculty of
Microbiology and Immunology, Department of
Graduate
6
Giske, Christian G. "Carbapenem resistance in Pseudomonas aeruginosa /." Stockholm, 2007. http://diss.kib.ki.se/2007/978-91-7357-080-0/.
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Henrichfreise, Beate. "Antibiotika-Multiresistenz bei Pseudomonas aeruginosa." [S.l.] : [s.n.], 2006. http://www.gbv.de/dms/bs/toc/517839636.pdf.
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Seabra, Rita A. M. "Immune modulation by Pseudomonas aeruginosa." Thesis, University of Nottingham, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.436865.
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Beatson, Scott. "Pseudomonas aeruginosa genomics and pathogenesis /." [St. Lucia, Qld.], 2002. http://www.library.uq.edu.au/pdfserve.php?image=thesisabs/absthe16848.pdf.
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Lasry, Judith. "Les Pseudomonas aeruginosa dans l'environnement." Paris 5, 1998. http://www.theses.fr/1998PA05P233.
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Kowalska, Karolina. "Formation of biofilm in Pseudomonas aeruginosa : molecular characterisation of the macromolecular complex Pel." Thesis, Aix-Marseille 2, 2011. http://www.theses.fr/2011AIX22003.
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Pseudomonas aeruginosa est une bactérie pathogène opportuniste qui peut développer deux modes d’infection. Les infections aigües sont associées à la production et à la sécrétion de toxines qui peuvent avoir un effet cytotoxique, alors que dans le contexte d’une infection chronique la bactérie a tendance à s’établir sous la forme d’un biofilm. Un biofilm est une population de microorganismes organisée en une communauté et attachée sur une surface. Cette surface peut être biotique ou abiotique. Les biofilms bactériens ont des caractéristiques intrinsèques qui les rendent plus résistants à des conditions environnementales difficiles (pH, oxygène, UV, flux, etc…). Dans le cadre d’une infection l’établissement du biofilm résulte aussi en une population bactérienne difficile à éliminer par le système immunitaire mais également résistante aux antibiotiques. Des déterminants moléculaires majeurs qui interviennent dans la formation du biofilm sont le flagelle, les pili de type IV et les fimbriae Cup pour l’attachement, ou bien encore les exopolysaccharides qui sont avec l’ADN des composants essentiels de la matrice extracellulaire du biofilm qui englobe la population bactérienne. Chez P. aeuginosa, si l’alginate est le polysaccharide majeur, il a été montré que des souches non-mucoides forment également des biofilms et que dans ce contexte la synthèse et la sécrétion du polysaccharide majeur de la matrice sont dépendantes des gènes pel. Mon travail a consisté dans l’étude de l’organisation structurale du système Pel et le contrôle de son activitéPseudomonas aeruginosa is a human opportunistic pathogen, and in the course of an infection can develop two lifestyles. One of them is involved in secretion of toxins and results in acute infection, and the other one causes chronic infections and is characterized by formation of a biofilm. A biofilm is a highly organized bacterial population, organized as a complex community that is attached to a surface. Bacterial biofilm shows higher resistance to different enviromental factors including physical factors, antimicrobials or host immune response. Major components taking part in biofilm formation are flagella, type IV pili, cup fimbriae as well as exopolysaccharides. The latter provides a protective matrix for the biofilm, together with proteins and DNA. In non-mucoid strains the major exopolysaccharide of biofilm matrix is synthetised and secreted by pel gene cluster. My work concentrates on the structural organisation of Pel polysaccharide secretion machinery, regulation of its activity as well as detailed characterisation of the Pel polysaccharide itself
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Wiehlmann, Lutz. "Sequenzspezifizierte Transposonmutagenese (STM) in Pseudomonas aeruginosa." [S.l. : s.n.], 2001. http://deposit.ddb.de/cgi-bin/dokserv?idn=96511211X.
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Juhas, Mario. "Global virulence regulators of Pseudomonas aeruginosa." [S.l.] : [s.n.], 2004. http://deposit.ddb.de/cgi-bin/dokserv?idn=974133221.
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Mettrick, Karla Adelle, and n/a. "Iron signalling pathways of Pseudomonas aeruginosa." University of Otago. Department of Biochemistry, 2008. http://adt.otago.ac.nz./public/adt-NZDU20081128.143145.
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The pathogenic bacterium Pseudomonas aeruginosa uses a variety of highly efficient chelating compounds (siderophores) to acquire sufficient iron for growth and virulence. These siderophores can either be endogenous or acquired from exogenous sources such as other bacteria or fungi. The transport of the endogenous siderophore pyoverdine activates a signal-transduction pathway that increases the synthesis of both the ferripyoverdine receptor protein (FpvA) and pyoverdine itself. Signal-transduction systems similar to this have three specific proteins involved: a receptor protein specific for one siderophore in the outer membrane, an anti-sigma factor in the cytoplasmic membrane and a sigma factor that activates gene expression in the cytoplasm.
The aim of the research presented in this thesis was to study the roles of the proteins in three different iron uptake and signalling pathways of P. aeruginosa. The substrates for each receptor protein were confirmed and the roles of each protein in the pathways were compared to the P. aeruginosa pyoverdine signalling pathway. The pyoverdine, desferrioxamine and ferrichrome transport pathways were studied to find whether interactions occur between them and if so, the mechanism(s) for that interaction. Furthermore, a technique for analysing gene expression of P. aeruginosa in sputum from the cystic fibrosis (CF) lung was developed. This technique was subsequently used to study the levels of iron responsive gene expression.
The receptor, sigma factor and anti-sigma factors were all found to have a role in the siderophore-induced expression of their own signalling pathway. The experimental data provide evidence of similarities in the roles of the sigma and receptor proteins within each pathway but different roles for the anti-sigma factors. In the absence of the cognate sigma factor or anti-sigma factor the expression of the desferrioxamine and ferrichrome receptors could not be upregulated. Without its cognate sigma factor fpvA could no longer be upregulated in the presence of pyoverdine. However, unlike the other systems, in the absence of the cognate anti-sigma factor, expression of fpvA was always observed. This is consistent with the anti-sigma factors being required for the activity of the cognate sigma factor in the ferrichrome and desferrioxamine signalling pathways but not the pyoverdine signalling pathway.
The siderophore signalling pathways were found to be upregulated in the presence of multiple siderophores, but generally to a lesser extent than if only one siderophore was available. This suggests that in the presence of multiple siderophores, P. aeruginosa uses all available iron chelators. The study of the role of the receptor, sigma factor and anti-sigma factor into these effects indicate sigma factor competition for RNA polymerase has a major role in the effects of multiple siderophores on pathways upregulation.
The gene expression studies of P. aeruginosa in sputum from the lungs of CF patients provided support for the hypothesis that the bacteria were growing in an environment where iron levels were sufficient for bacterial growth, but not storage of iron. The expression of the sigma factor gene pvdS that is required for pyoverdine synthesis was studied because expression of this gene is a sensitive reporter of intracellular iron levels. It was found to be downregulated in bacteria in sputum compared to laboratory grown bacteria. This result suggests the bacteria are inhabiting a more iron-replete environment within the lung. This finding advances our understanding of the CF lung environment and the impact it has on P. aeruginosa infection. This knowledge has medical implications for the development of novel therapies to combat P. aeruginosa infection.
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Léger, Jean-François. "Effects of chloramphenicol on Pseudomonas aeruginosa." Thesis, McGill University, 1991. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=60549.
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The characteristics of the effects of chloramphenicol on Pseudomonas aeruginosa were examined. Resistant strains were easily isolated following a single passage in chloramphenicol at 150 $ mu$g/ml to 500 $ mu$g/ml. Drug detoxification or altered sensitivity of the target site could not be the mechanism of resistance. This resistance to chloramphenicol was correlated with the addition of an outer membrane protein with a molecular weight of 49 kDa and the loss of two outer membrane proteins, one with the molecular weight of 19 kDa and the other of about 10 kDa. The highly specific requirement of the resistant strains for Ca$ sp{2+}$, Mg$ sp{2+}$, Mn$ sp{2+}$ or Sr$ sp{2+}$ described by Irvin and Ingram (1982) was confirmed by the observation that the outer membrane of the resistant cells contained twice as much Mg$ sp{2+}$ cation as the sensitive cells. Many other experiments designed to observe the effects of chloramphenicol on the outer membrane of P. aeruginosa failed. It was concluded that the observations made in this study strongly suggested a "re-structuring" of the outer membrane of P. aeruginosa, rendering the resistant cells more impermeable to chloramphenicol.
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Wang, Chan-Ju. "Characterisation of azoreductases form pseudomonas aeruginosa." Thesis, University of Oxford, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.531806.
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Hughes, M. A. "Transfer RNA genes in Pseudomonas aeruginosa." Thesis, University of Liverpool, 1986. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.370854.
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Balasubramanian, Deepak. "Pseudomonas Aeruginosa AmpR Transcriptional Regulatory Network." FIU Digital Commons, 2013. http://digitalcommons.fiu.edu/etd/863.
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In Enterobacteriaceae, the transcriptional regulator AmpR, a member of the LysR family, regulates the expression of a chromosomal β-lactamase AmpC. The regulatory repertoire of AmpR is broader in Pseudomonas aeruginosa, an opportunistic pathogen responsible for numerous acute and chronic infections including cystic fibrosis. Previous studies showed that in addition to regulating ampC, P. aeruginosa AmpR regulates the sigma factor AlgT/U and production of some quorum sensing (QS)-regulated virulence factors. In order to better understand the ampR regulon, the transcriptional profiles generated using DNA microarrays and RNA-Seq of the prototypic P. aeruginosa PAO1 strain with its isogenic ampR deletion mutant, PAO∆ampR were analyzed. Transcriptome analysis demonstrates that the AmpR regulon is much more extensive than previously thought influencing the differential expression of over 500 genes. In addition to regulating resistance to β-lactam antibiotics via AmpC, AmpR also regulates non-β-lactam antibiotic resistance by modulating the MexEF-OprN efflux pump. Virulence mechanisms including biofilm formation, QS-regulated acute virulence, and diverse physiological processes such as oxidative stress response, heat-shock response and iron uptake are AmpR-regulated. Real-time PCR and phenotypic assays confirmed the transcriptome data. Further, Caenorhabditis elegans model demonstrates that a functional AmpR is required for full pathogenicity of P. aeruginosa. AmpR, a member of the core genome, also regulates genes in the regions of genome plasticity that are acquired by horizontal gene transfer. The extensive AmpR regulon included other transcriptional regulators and sigma factors, accounting for the extensive AmpR regulon. Gene expression studies demonstrate AmpR-dependent expression of the QS master regulator LasR that controls expression of many virulence factors. Using a chromosomally tagged AmpR, ChIP-Seq studies show direct AmpR binding to the lasR promoter. The data demonstrates that AmpR functions as a global regulator in P. aeruginosa and is a positive regulator of acute virulence while negatively regulating chronic infection phenotypes. In summary, my dissertation sheds light on the complex regulatory circuit in P. aeruginosa to provide a better understanding of the bacterial response to antibiotics and how the organism coordinately regulates a myriad of virulence factors.
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Chen, Chun Chiang. "Rhamnolipid Production with Denitrifying Pseudomonas Aeruginosa." University of Akron / OhioLINK, 2006. http://rave.ohiolink.edu/etdc/view?acc_num=akron1236689824.
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Moore, Matthew Phillip. "Comparative genomics of Pseudomonas aeruginosa populations." Thesis, University of Liverpool, 2017. http://livrepository.liverpool.ac.uk/3021281/.
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Pseudomonas aeruginosa causes a wide range of infections, is often associated with antimicrobial resistance and is the primary cause of chronic lung infection in cystic fibrosis (CF) and the overall morbidity and mortality associated with the disease. As P. aeruginosa is an environmental bacterium that opportunistically infects CF patients most infecting lineages are distinct. The determination of common adaptive routes during infection is further complicated by within-lineage heterogeneity, multi-lineage infections and the emergence of transmissible strains. In order to better understand the genetic basis of varying pathogenicity, a diverse dataset of 1,407 P. aeruginosa genomes from the environment, non-CF bronchiectasis and from CF samples was analysed. Detailed analysis of mutations potentially associated with the CF lung environment was conducted by comparison of the genomes from CF and environmental isolates and by including a geographically and historically diverse panel of a transmissible lineage, the Liverpool Epidemic Strain (LES). Sequencing of isolates from non-CF bronchiectasis showed for the first time the commonality in adaptive routes in non-CF chronic lung diseases and the utility of genome sequencing to infer within and between host diversity and population structure. Many antibiotic resistance genes (ARGs) in all samples were observed to be adaptive and a sample of genomes from hospital isolates from Thailand was used to assess international multi-drug resistance lineages with mobile genetic element associated ARGs. This study has shown, by analysis of diverse datasets, how genome sequencing can reveal the genetic basis of phenotypic heterogeneity and subsequent varying patient outcomes with this diverse infection.
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Stacey, Sean D. "Regulating rsmA Expression in Pseudomonas aeruginosa." Digital Commons @ East Tennessee State University, 2013. https://dc.etsu.edu/etd/1232.
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Pseudomonas aeruginosa, a Gram-negative bacillus, commonly infects immunocompromised individuals and uses a variety of virulence factors to persist in these hosts. The posttranscriptional regulator, RsmA, plays a role in the expression of many virulence factors in P. aeruginosa. RsmA up regulates virulence factors used in colonizing hosts. However, regulation of rsmA is not well elucidated. Transposon mutagenesis was performed on P. aeruginosa containing a transcriptional rsmA-lacZ fusion to answer this question. Mutants were screened via β-galactosidase assay and transposon insertions identified via arbitrary PCR. A probable MFS transporter, we named mtpX, was one significant transposon mutant identified. A ΔmtpX mutant containing the rsmA-lacZ transcriptional fusion was constructed to confirm our results. Further analysis of rsmA, looking at RNA and protein levels, revealed varying results in nonmucoid versus mucoid backgrounds. Phenotypic assays were performed to characterize this unknown transporter and develop a putative mechanism as to how MtpX affects rsmA expression.
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O'Toole, Ann Marie. "Thermal deactivation of Pseudomonas aeruginosa biofilms." Thesis, University of Iowa, 2015. https://ir.uiowa.edu/etd/1715.
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Bacterial biofilm infection is a common (~ 2 to 4%) complication for recipients of surgically implanted medical devices. Due to the tremendous increase in antibiotic resistance when these bacteria enter the biofilm phenotype, present treatment requires explantation and replacement of the device, often with multiple surgeries and always with much longer patient recovery time. The specific objective of this study was to quantify the degree of biofilm deactivation from exposure to thermal shock for varying temperature and time durations. While extreme temperature (>150˚C) is routinely used to sterilize (e.g. autoclaves), such temperatures have a severe cost within the body. Despite extensive studies on thermal deactivation of bacteria in the planktonic phenotype over a wide range of temperatures (e.g., pasteurization protocols), surprisingly little is known about the thermal deactivation of biofilms except under extreme conditions. Here, the deactivation of Pseudomonas aeruginosa biofilms is reported. These biofilms were cultured at 37°C for 24 hours in a drip-flow reactor and subjected to heat shocks on the range of 50°C to 80°C for durations of 1 to 30 minutes. Heat shocks were delivered by immersion in thermostatted media for the prescribed time and the resulting concentration of colony forming units (CFU/mL) were quantified using direct enumeration. Up to 6.6 orders of magnitude reduction in CFU concentration was observed, indicating that thermal deactivation is a reasonable approach to biofilm mitigation. Integrating this approach with a magnetic nanoparticle implant coating will result in an innovative treatment for implant infections in situ without explantation or device replacement.
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Cacci, Luciana Camila. "Estudo epidemiologico-molecular das infecções por pseudomonas aeruginosa resistente ao imipenem em pacientes hospitalizados." [s.n.], 2007. http://repositorio.unicamp.br/jspui/handle/REPOSIP/310665.
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Orientador: Marcelo de Carvalho RamosDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas
Made available in DSpace on 2018-08-10T01:23:41Z (GMT). No. of bitstreams: 1 Cacci_LucianaCamila_M.pdf: 1733904 bytes, checksum: 2616f843895d251e8e236cdd873fe436 (MD5) Previous issue date: 2007
Resumo: Introdução: As metalo beta-lactamases, presentes em Pseudomonas aeruginosa assim como em diversos microrganismos Gram negativos, são grande ameaça ao tratamento dos pacientes portadores de infecções causadas por este microrganismo. Objetivos: A produção de MBLs e a relação genética foram investigadas em isolados de P. aeruginosa resistentes ao Imipenem, recuperados de infecções hospitalares. Descrição do estudo: Estudo restropectivo em uma amostra de microrganismos. O estudo foi conduzido em dois hospitais universitários, em Campinas. Todos os isolados de P. aeruginosa resistentes ao Imipenem foram coletados de pacientes hospitalizados no período de Março de 2000 a Dezembro de 2004. Métodos: O método da disco-difusão foi utilizado para confirmar a resistência ao Imipenem. O E-test MBL@ foi feito para verificar a produção de MBLs e a Concentração Inibitória Mínima (CIM) do Imipenem. Os ftagmentos das seqüências dos genes blaIMP-I, blaVIM-I, blaVIM-2 e blaSPM-I foram amplificados. Resultados: Cento e vinte e oito isolados resistentes ao Imipenem foram coletados durante o período do estudo. A maioria dos isolados exibiu CIM do Imipenem maior ou igual a 256 ug/mL. A análise por macrorestrição com a enzima SpeI através do Pulsed Field Gel Electrophoresis (PFGE) mostrou um polimorfismo significativo. Apenas 15 cepas puderam ser distribuídas em sete "clusters", seis com dois isolados e um com três isolados. Noventa e oito isolados resistentes ao Imipenem foram triados para a pesquisa da produção de MBL. Setenta isolados apresentaram produção de MBL e o fragmento do gene blaSPM-l pôde ser amplificado em 12 isolados. Nas cepas restantes nenhum outro tipo de MBL referente aos genes blaIMP-l, blaVIM-I, blaVIM-2 foi encontrado. Conclusão: A disseminação de cepas de P. aeruginosa produtoras de MEL genotipicamente heterogêneas foi documentada nos hospitais estudados. Apenas a metalo beta-lactamase SPM-1 foi encontrada entre essas cepas
Abstract: Objective: Genetic relatedness and Metallo-lactamase production was investigated in Imipenem resistant Pseudomonas aeruginosa recovered from hospital acquired infections. Design: Descriptive study in a convenient sample of organisms. Setting: Two 400-bed tertiary care teaching hospitals, in Campinas, Brazil. All Imipenem resistant P. aeruginosa, recovered from March, 2000 through December 2004 from hospitalized patients were collected Methods: Disk diffusion tests were used to confirm Imipenem resistance. E-test MBL@ was done to check for MBL production, ando Imipenem MIC's blasPM-l, .blaIMP-l, blaVIM-l and blaVIM-2 sequences were amplified. Results: A sample of 128 Imipenem resistant P. aeruginosa isolates was collected during the study period. Most isolates exhibited Imipenem MIC's > 256 ug/mL. Macrorestriction analysis (Spel) using pulsed field gel electrophoresis (PFGE) showed a substantial polymorphism. Only 15 strains could be allocated to seven clusters, six with two isolates and one with three isolates. Ninety-eight Imipenem resistant isolates were screened for MBL production. Seventy isolates showed MBL production, and blasPM-l sequence could be amplified from 12 isolates. In the remaining strains no other MBL-type from blaIMP-l, blaVIM-l and blaVIM-2 investigated in the study was demonstrated. Conclusion: Dissemination of MBL producing-genotypically heterogenous Pseudomonas aeruginosa strains was documented in the hospitals studied. Only SPM-l metallo-_-lactamase was found among these strains
Mestrado
Ciencias Basicas
Mestre em Clinica Medica
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Ledgham, Fouzia. "La régulation des facteurs de virulence chez Pseudomonas aeruginosa : Quorum Sensing et synthèse d'alginate." Aix-Marseille 2, 2001. http://www.theses.fr/2001AIX22034.
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Damron, Frederick H. "Regulation of alginate production of Pseudomonas aeruginosa." [Huntington, WV : Marshall University Libraries], 2009. http://www.marshall.edu/etd/descript.asp?ref=999.
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Chemani, Chanez, and Régis Matran. "Les lectines : nouveaux déterminants de la pathogénicité de Pseudomonas aeruginosa au cours de l'infection pulmonaire : vers de nouvelles thérapeutiques." Lille 2, 2009. http://www.theses.fr/2009LIL2S020.
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Pseudomonas aeruginosa est un pathogène fréquemment impliqué dans les infections pulmonaires aiguës et chroniques, notamment chez les patients atteints de mucoviscidose. L'adhésion de cette bactérie est principalement médiée par des lectines qui reconnaissent spécifiquement des structures glycaniques de l'hôte. Dans ce travail, nous avons évalué le rôle respectif de deux lectines LecA et LecB dans la pathogénicité de P. Aeruginosa ainsi que les effets de sucres inhibiteurs spécifiques de celles-ci dans un modèle de pneumonie aiguë chez la souris et sur un modèle cellulaire. En comparant une souche parentale à ses deux mutants isogéniques (mutant lecA et lecB), nous avons montré que ces deux lectines jouent un rôle majeur dans l'adhésion de P. Aeruginosa aux cellules A549 et sont associées à la sévérité de la lésion pulmonaire in vivo. L'inhibition de ces deux lectines diminue significativement la cytotoxicité in vitro et augmente la clairance bactérienne pulmonaire in vivo. Nous avons ensuite évalué la contribution des deux lectines à l'initiation et l'installation de l'infection chronique. Dans un modèle murin de pneumonie chronique, nous avons observé une mortalité moins importante avec les mutants lecA et lecB et une diminution de la de la charge bactérienne pulmonaire au 4ème et 7ème jour suivant l'infection. L'analyse de l'expression des gènes de virulence au cours de l'infection chronique montre une surexpression d'exoS et de lasI chez les mutants lectines par rapport à la souche parentale. Les capacités thérapeutiques d'inhibiteurs glycomimétiques ayant un potentiel d'inhibition 7 à 10 fois supérieur aux ligands naturels ont été évaluées. Ces molécules de synthèse diminuent significativement l'adhésion de P. Aeruginosa in vitro et montrent un effet protecteur vis-à-vis de la lésion pulmonaire in vivo. En conclusion, nous avons montré que les lectines jouent un rôle majeur dans la pathogénicité de P. Aeruginosa. L'inhibition des lectines par des sucres compétiteurs pourrait constituer une nouvelle piste thérapeutique dans l'infection à P. Aeruginosa.
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Douthett, Rebecca L. "Enhancement of the humoral immune response to Pseudomonas aeruginosa." The Ohio State University, 2005. http://rave.ohiolink.edu/etdc/view?acc_num=osu1126903068.
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Klebensberger, Janosch. "Detergent-induced cell aggregation in Pseudomonas aeruginosa." [S.l. : s.n.], 2007. http://nbn-resolving.de/urn:nbn:de:bsz:352-opus-26614.
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Dumenil, Sylvie. "Influence de concentrations subinhibitrices d'amikacine sur l'activité élastolytique de"Pseudomonas aeruginosa" IFO 3455." Paris 5, 1991. http://www.theses.fr/1991PA05P101.
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Treepong, Panisa. "Bioinformatic analysis of the genomes of epidemic pseudomonas aeruginosa." Thesis, Bourgogne Franche-Comté, 2017. http://www.theses.fr/2017UBFCD065/document.
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Le Pseudomonas aeruginosa est un pathogène nosocomial majeur. Le clone ST235 est le plus prévalent des clones internationaux dits à hautris que. Ce clone est très fréquemment multi résistant aux antibiotiques, ce qui complique la prise en charge des infections dont il est à l’origine.Malgré son importance clinique, la base moléculaire Du succès du clone ST235 n’est pas comprise.Dans ce travail, nous avons cherché à comprendre l’origine spacio temporelle de ce clone et les bases moléculaires de son succès. A l’aide d’outils bio informatiques existants ,nous avons trouvé que le clone ST235 a émergé en Europe en 1984 et que tous les isolates ST235 produisent l’exotoxine ExoU. Nous avons également identifié 22 gènes Contigus spécifiques de ce clone et impliqués dans l’efflux transmembranaire, dans le traitement de l’ADN et dans la transformation bactérienne. Cette combinaison unique de gènes a pu contribuer à la gravité des infections dues à ce clone et à sa capacité à acquérir des gènes de résistance aux antibiotiques. Ainsi, la diffusion mondiale de ce clone a probablement été favorisée par l’utilisation extensive des fluoroquinolones, puis il est de venu localement résistant aux amino glycosides, aux β-lactamines, et aux carbapénèmes par mutation et acquisition d’éléments de résistance. Nous avons majoritairement utilisé des outils existants,mais avons découvert que les programmes de détection des séquences d’insertions (IS, ayant un rôle important dans l’évolution des génomes bactériens) ne sont pas adaptés aux données dont nous disposions. Nous avons ainsi mis au point un outil (appelé panISa) qui détecte de façon précise et sensible les IS à partir de données brutes de séquençage de génomes bactériensPseudomonas aeruginosa is a major nosocomial pathogen with ST235 being the most prevalent of the so-called ‘international’ or ‘high-risk’ clones. This clone is associated with poor clinical outcomes in part due to multi- and high-level antibiotic resistance. Despite its clinical importance, the molecular basis for the success of the ST235 clone is poorly understood. Thus this thesis aimed to understand the origin of ST235 and the molecular basis for its success, including the design of bioinformatics tools for finding insertion sequences (IS) of bacterial genomes.To fulfill these objectives, this thesis was divided into 2 parts.First, the genomes of 79 P. aeruginosa ST235 isolates collected worldwide over a 27-year period were examined. A phylogenetic network was built using Hamming distance-based method, namely the NeighborNet. Then we have found the Time to the Most Recent Common Ancestor (TMRCA) by applying a Bayesian approach. Additionally, we have identified antibiotic resistance determinants, CRISPR-Cas systems, and ST235-specific genes profiles. The results suggested that the ST235 sublineage emerged in Europe around 1984, coinciding with the introduction of fluoroquinolones as an antipseudomonal treatment. The ST235 sublineage seemingly spreads from Europe via two independent clones. ST235 isolates then appeared to acquire resistance determinants to aminoglycosides, β-lactams, and carbapenems locally. Additionally, all the ST235 genomes contained the exoU-encoded exotoxin and identified 22 ST235-specific genes clustering in blocks and implicated in transmembrane efflux, DNA processing and bacterial transformation. These unique genes may have contributed to the poor outcome associated with P. aeruginosa ST235 infections and increased the ability of this international clone to acquire mobile resistance elements.The second part was to design a new Insertion Sequence (IS) searching tool on next-generation sequencing data, named panISa. This tool identifies the IS position, direct target repeats (DR) and inverted repeats (IR) from short read data (.bam/.sam) by investigating only the reference genome (without any IS database). To validate our proposal, we used simulated reads from 5 species: Escherichia coli, Mycobacterium tuberculosis, Pseudomonas aeruginosa, Staphylococcus aureus, and Vibrio cholerae with 30 random ISs. The experiment set is constituted by reads of various lengths (100, 150, and 300 nucleotides) and coverage of simulated reads at 20x, 40x, 60x, 80x, and 100x. We performed sensitivity and precision analyses to evaluate panISa and found that the sensitivity of IS position is not significantly different when the read length is changed, while the modifications become significant depending on species and read coverage. When focusing on the different read coverage, we found a significant difference only at 20x. For the other situations (40x-100x) we obtained a very good mean of sensitivity equal to 98% (95%CI: 97.9%-98.2%). Similarly, the mean of DR sensitivity of DR identification is high: 99.98% (95%CI: 99.957%-99.998%), but the mean of IR sensitivity is 73.99% (95%CI: 71.162%-76.826%), which should be improved. Focusing on precision instead of sensibility, the precision of IS position is significantly different when changing the species, read coverage, or read length. However, the mean of each precision value is larger than 95%, which is very good.In conclusion, P. aeruginosa ST235 (i) has become prevalent across the globe potentially due to the selective pressure of fluoroquinolones and (ii) readily became resistant to aminoglycosides, β-lactams, and carbapenems through mutation and acquisition of resistance elements among local populations. Concerning the second point, our panISa proposal is a sensitive and highly precise tool for identifying insertion sequences from short reads of bacterial data, which will be useful to study the epidemiology or bacterial evolution
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Head, Nathaniel Edwards. "Regulation of biofilm formation of pseudomonas aeruginosa." Huntington, WV : [Marshall University Libraries], 2006. http://www.marshall.edu/etd/descript.asp?ref=663.
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Limpert, Anna Silke. "Functional genome analysis in Pseudomonas aeruginosa SG17M." [S.l.] : [s.n.], 2005. http://deposit.ddb.de/cgi-bin/dokserv?idn=976191474.
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Fakhimi, Tatiana. "Sekundära metaboliters antibakteriella effekt mot Pseudomonas aeruginosa." Thesis, Umeå universitet, Farmakologi, 2018. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-146099.
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Handfield, Martin. "Expression génique in vivo chez Pseudomonas aeruginosa." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk3/ftp04/nq25421.pdf.
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Meldrum, Allison J. "Regulation of pyoverdine biosynthesis in Pseudomonas aeruginosa." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk2/ftp01/MQ37969.pdf.
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Purevdorj-Gage, Boloroo. "Pseudomonas aeruginosa Biofilm Structure, Behavior and Hydrodynamics." Thesis, Montana State University, 2004. http://etd.lib.montana.edu/etd/2004/purevdorj-gage/Purevdorj-GageB1204.pdf.
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Biofilm formation by bacterial pathogens is an important factor in the progression and treatment of many infectious diseases. Biofilm structural development is a dynamic process dependent on many cellular and environmental parameters including Quorum Sensing (QS) and hydrodynamics. Since QS is dependent on a threshold autoinducer concentration, it was hypothesized that the flow dynamics in the bulk fluid surrounding the biofilm would play an important role in expression of QS and the genes that are under its control. In order to investigate the relative contribution of hydrodynamics and QS on biofilm development, biofilms were grown from wild type Pseudomonas aeruginosa PAO1 and the cell signaling lasI mutant PAO1-JP1 under laminar and turbulent flows. When morphology of the biofilms were quantified using Image Structure Analyzer (ISA) software, a multivariate analysis demonstrated that both QS and hydrodynamics influenced biofilm structure, suggesting that QS was not required for biofilm development but affected structural heterogeneity in biofilms. GFP reporter based gene expression analysis of QS regulated lasB (coding for elastase) expression during biofilm development in laminar flow further supported these results. Detachment has been recognized as another factor that may define structural morphology of biofilms. Under flow conditions hollow biofilm clusters were formed as a result of active detachment process, termed as "seeding dispersal". A differentiation of a "seeding" microcolony into an interior motile, swarming, phenotype and a non-motile surrounding, "wall phenotype" formed as a prelude to the dispersal process in which the interior cells swarmed out of the microcolony from local break out points and spread over the wall of the flow cell. A critical microcolony diameter of approximately 100 um was required for differentiation suggesting that regulation was related to cell density and mass transfer conditions. It was found that rhamnolipid (rhlA-) biosurfactant was not required and QS system (PAO1-JP2) was shown to be important in this process, possibly by sensing nutrient limitation within the biofilm microcolonies. These results strengthen a current view of multi-cellularity and coordinated behavior in prokaryotes as well as a dynamic network of overlapping pathways and cellular mechanisms that act on biofilm development in a complex interrelated manner.
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Xu, Ruifang. "Spatial Growth Patterns of Pseudomonas aeruginosa Biofilms." Thesis, Montana State University, 2004. http://etd.lib.montana.edu/etd/2004/xu/XuR0805.pdf.
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Biofilms are less susceptible to antimicrobial action compared to their planktonic counterparts. The protective mechanisms are not fully understood. Physiological heterogeneity within biofilms is thought to contribute to the low susceptibility and was therefore studied. Expression of green fluorescent protein (GFP), induction of alkaline phosphatase (APase) by phosphate starvation, and the cell viability assay using the LIVE/DEAD BacLight bacterial viability stain were performed to visualize the spatial patterns of growth and viability within 5-d-old Pseudomonas aeruginosa biofilms. The capillary reactor and the drip-flow reactor were employed to obtain biofilms of a range of thickness. Biofilms cultivated in the capillary reactor were usually thinner than those grown in the low-shear drip-flow reactor. The former were examined by in situ confocal scanning laser microscopy (CSLM) whereas the latter were cryoembedded and cryosectioned prior to conventional fluorescence microscopic observation. P. aeruginosa PAO1 with the plasmid pAB1 carrying an inducible, stable gfp was used to identify zones of active protein synthesis. The induction of gfp proved suitable for the visualization of spatial growth patterns within biofilms. Greater GFP activity was evident at the surface of clusters and was not as bright in their centers after induction. Activity appeared more uniform in smaller clusters and less uniform in larger clusters. The APase activity induced by phosphate starvation showed a sharply delineated band of active APase synthesizing cells close to the biofilm-bulk fluid interface and some local APase synthetic activity in the depth of the biofilm. The results of biofilm viability staining using the LIVE/DEAD BacLight bacterial viability kit turned out to be puzzling and cast doubt on the methodological validity of applying the LIVE/DEAD BacLight bacterial viability staining method to P. aeruginosa biofilms. The findings of the spatial growth patterns illustrate the physiological heterogeneity that is present in these biofilms. Such variation in the metabolic activity probably contributes to the reduced susceptibility of these biofilms to antimicrobial agents.
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Fu, Yinan. "Structure and dynamics of Pseudomonas aeruginosa ICP." Thesis, University of Glasgow, 2009. http://theses.gla.ac.uk/723/.
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Pseudomonas aeruginosa inhibitor of cysteine peptidases (PA-ICP) is a potent protein inhibitor of papain-like cysteine peptidases (CPs) identified in Pseudomonas aeruginosa, an opportunistic pathogenic bacteria that can cause severe infections in human. It belongs to the newly characterized natural CP inhibitors of the I42 family, designated the ICP family. The members of this family are present in some protozoa and bacterial pathogens. They can inhibit both parasite and mammalian CPs with high affinity and specificity. Whether the main biological function of the proteins in the pathogens is to regulate the hydrolytic activity of the organisms’ endogenous CPs or exogenous CPs so as to facilitate the pathogens’ invasion or survival is still under investigation. Although Pseudomonas aeruginosa contains a CP inhibitor, no CP genes are found in its genome, suggesting that the targets of PA-ICP may be exogenous. This hypothesis is supported by the presence of a putative secretion signal peptide at the N-terminus of PA-ICP which may be involved in exporting the protein to target exogenous CPs. In order to shed light on the biological function and inhibitory specificity of PA-ICP, the structure and backbone dynamics of this protein were characterised using NMR spectroscopy. In this project, the inhibitory activity of PA-ICP to a range of mammalian model CPs was also studied. Like its previously studied homologs, PA-ICP adopts an immunoglobulin fold comprised of seven β-strands. Three highly conserved sequence motifs located in mobile loop regions form the CP binding site. The inhibitor exhibits higher affinity toward the mammalian CP cathepsin L than cathepsins H and B. Homology modelling of the PA-ICP-cathspin L interaction based on the crystal structure of the chgasin-cathpsin L complex shows that PA-ICP may inhibit the peptidases by blocking the enzyme’s active site and that the interactions between chagasin and CPs may be conserved in PA-ICP-peptidase complexes. The specificity of the inhibitors may be determined by the relative flexibility of the loops bearing the binding site motifs and the electrostatic properties of certain residues near the binding sites.
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Worrall, Kathryn E. "The role of MvaT in Pseudomonas aeruginosa." Thesis, University of Nottingham, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.430524.
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Gifford, Danna R. "Population genetics of rifampicin-resistant Pseudomonas aeruginosa." Thesis, University of Oxford, 2014. http://ora.ox.ac.uk/objects/uuid:044b9258-4f10-4e77-9ff6-aa4035cec33b.
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Antibiotic resistance is generally associated with a cost in terms of reduced competitive fitness in the absence of antibiotics. Despite this 'cost of resistance', the cessation of antibiotic treatment does not result in significant reductions in the prevalence of resistance. The maintenance of resistance, in spite of the costs, has been attributed to the rarity of reversion mutations, relative to compensatory mutations at other loci in the genome. However, the large size of bacteria populations, and the potential for migration, suggest that reversion mutations should occasionally be introduced to resistant populations. In this thesis, I show that additional mechanisms can prevent fixation of reversion mutations even if they do occur. Using an experimental evolution approach, with rifampicin resistance in Pseudomonas aeruginosa as a model system, I measured the costs of resistance in several environments and followed the adaptive dynamics of resistant populations where a sensitive lineage had invaded by migration. The results suggest that several additional mechanisms contribute to the maintenance of antibiotic resistance. Most rifampicin resistance mutations are not unconditionally costly in all environments, suggesting that migration between environments could maintain a resistant reservoir population. In environments where resistance is initially costly, the fixation of a revertant is not guaranteed, even if introduced through migration. Revertant fixation was impeded or prevented by clonal interference from adaptation in the resistant strain. Revertants that did successfully replace the resistant strain were forced to adapt to do so. Contrary to assumptions in the existing literature, fitness in the resistant strains was not recovered by general compensatory mutations, but instead by adaptive mutations specific to the environment. The data challenge several assumptions about the maintenance of antibiotic resistance: that resistance mutations are always costly, that the rarity of back mutations prevents the reversion of resistance, and that resistant strains recover fitness by compensatory mutations.
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Crescente, Vincenzo. "Azoreductases : genes and proteins in Pseudomonas aeruginosa." Thesis, Kingston University, 2015. http://eprints.kingston.ac.uk/36328/.
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Pseudomonas aeruginosa is one of the primary causes of opportunistic infections in humans and it is associated with both acute and chronic infections in immunocompromised individuals. This bacterium is extremely resistant to many antibiotics, making the treatments against this pathogen often unsuccessful. Azoreductases, a family of enzymes involved in the reduction of azo compounds and quinones, are found in many bacterial species including P. aeruginosa. Although the enzymatic activity of three of these enzymes has been extensively characterized, their physiological role remains unclear. In this study, the enzymatic activity as well as the effect on physiological processes such as swarming motility, biofilm formation and antibiotic resistance of known and putative azoreductase proteins from P. aeruginosa PAO1 have been investigated. Five putative azoreductase genes from P. aeruginosa PAO1 (pa0949, pa1204, pa2280, pa2580 and pa4975) were cloned and four of these (pa0949, pa1204, pa2280 and pa2580) were over expressed in E. coli strains. Recombinant proteins were purified and biochemically characterized showing the presence of FAD bound to PA1204 and PA2580 proteins. Enzymatic reaction conditions were established for each protein by determining the preferred cofactor and reductant (flavin and NAD(P)H) used by each protein. Higher reduction rates were obtained using FAD for PA1204 and PA2580, and FMN for PA0949 and PA2280, whereas NADPH was always the preferred reductant for all of the enzymes tested. Substrate specificity studies performed with azo compounds and quinones showed that PA1204, PA2280 and PA2580 recombinant proteins can reduce both classes of substrate, with higher reduction rates with quinones, whereas the recombinant PA0949 protein showed to reduce only quinone substrates. Investigation of the role of azoreductase genes on P. aeruginosa PAO1 motility, biofilm formation and antibiotic resistance was conducted using single gene transposon mutants for each of the genes paazor1, paazor2, paazor3, pa0949, pa1204, pa2280, pa2580 and pa4975. Motility analysis demonstrated greater swarming for all mutants tested compared with wild type, although both wild type and mutants showed the same growth rate. Similarly, all mutants showed higher biofilm production compared with the wild type in a short term period (24 hours), whereas no differences were observed after a longer biofilm production period (48 hours). The MICs and MBCs of several antibiotics were determined, showing that, in presence of fluoroquinolones, P. aeruginosa PAO1 azoreductase mutants exhibit higher growth inhibition (up to 127-fold) and reduced survival (up to 7 fold) compared with wild type. This suggests that azoreductase genes (or gene products) may be involved in the P. aeruginosa PAO1 resistance to antibiotic treatments, and in particular to fluoroquinolones. The findings prove that the proteins PA0949, PA1204, PA2280 and PA2580 have similar features and enzymatic functions to the already characterized paAzoR1-3 from P. aeruginosa PAO1. Therefore, these can be included in the family of azo- and quinone- oxidoreductase enzymes. The data presented here on the antibiotic resistance strongly suggest a role for azoreductase gene products in antimicrobial resistance in P. aeruginosa. These original findings provide a springboard for further investigation of azoreductases as novel targets for antimicrobial agents for this pathogen.
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Davies, Emily. "The role of prophages in Pseudomonas aeruginosa." Thesis, University of Liverpool, 2015. http://livrepository.liverpool.ac.uk/2034639/.
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Pseudomonas aeruginosa is a common opportunistic respiratory pathogen of individuals with cystic fibrosis (CF), capable of establishing chronic infections in which the bacterial population undergoes extensive phenotypic and genetic diversification. The Liverpool Epidemic Strain (LES) is a widespread hypervirulent and transmissible strain that is capable of superinfection and is linked to increased morbidity and mortality, relative to other P. aeruginosa strains. The LES has six prophages (LESφ1-6) within its genome, of which three are essential to the competitiveness of this strain. Temperate bacteriophages are incredibly common in bacterial pathogens and can contribute to bacterial fitness and virulence through the carriage of additional genes or modification of existing bacterial genes, lysis of competitors, or by conferring resistance to phage superinfection. Furthermore, the LES phages are detected at high levels in the CF lungs and have been implicated in controlling bacterial densities. The aims of this study were to (i) further characterise the LES phages and their induction, (ii) determine the extent to which the LES phages contribute to bacterial phenotypic and (iii) genetic diversification and (iv) determine how the LES phages affect host competitiveness, using a variety of in vitro and in vivo infection models. LES phages are continuously produced by spontaneous lysis and this study found that environmental factors that are common to the CF lung, such as oxidative stress, pharmaceutical chelating agents and antibiotics, can alter phage production by clinical LES isolates. Characterisation of the phages highlighted differences between the phages with regards to their lytic cycles and ability to propagate in different environments. P. aeruginosa undergoes extensive phenotypic diversification in an artificial sputum model (ASM) of infection, similar to that observed in chronic CF infections. Hypermutability, loss of motility and auxotrophy were phenotypes observed in bacteria evolved for approximately 240 bacterial generations in ASM in the presence and absence of the LES phages. However, the LES phages accelerated this process; loss of twitching motility occured earlier in populations evolved in the presence of phages. Sequencing of evolved populations revealed a high level of genetic diversification, with genes involved in motility, quorum sensing and genetic regulation experiencing loss of function mutations in parallel populations. In phage treated populations, LESφ4 had disruptively integrated into motility and quorum sensing genes, suggesting that temperate phages can provide an alternative (and quicker) route to adaptation. LES prophage carriage is important for bacterial competitiveness; PAO1 LES Phage Lysogens (PLPLs) successfully invaded a phage-susceptible population in vitro from when initially rare. Strain invasiveness was dependent on the LES prophage; LESφ4 lysogens were more invasive than PLPLφ2 or PLPLφ3, whereas carriage of all three prophages accelerated bacterial invasion. PLPLφtriple could also invade a susceptible competitor population in a rat model of chronic lung infection, although not as successfully as in vitro. These data suggest that prophage carriage is important for LES competitiveness and that phage-mediated lysis of phage-susceptible competitors may explain why LES is adept at superinfection. The study indicates that the LES phages are important drivers of bacterial diversification and evolution and confer a competitive advantage to their bacterial host. This may help explain why the LES is so successful, and the high prevalence of polylysogeny in bacterial pathogens.
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Bhamra, Amrat. "Pathogenesis of Pseudomonas aeruginosa in leukaemic patients." Thesis, University of Surrey, 1993. http://epubs.surrey.ac.uk/843880/.
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The incidence of production and biological activity of various extracellular factors of Pseudomonas aeruginosa from leukaemic and non-leukaemic patients were investigated. A panel of 157 isolates was constructed from blood of leukaemic (Group I), and other body sites of the same patients (Group II), and from various specimens of non-leukaemic patients (Group III). Most strains produced pyocyanin. Cross infection between patients was not a significant problem. The incidence of protease production was high (94%) and there was no significant quantitative difference between strains. Protease production was inhibited by plasma a2 macroglobulin but urine was not inhibitory for the enzyme. Protease activity was demonstrated with fibronectin by a specific ELISA. Complement degradation, particularly C3, was also demonstrated by gel electrophoresis and immunoglobulins IgG, M and A were destroyed by HPLC purified alkaline protease. The molecular weight of alkaline protease was found to be 36 kd and that of elastase to be 22 kd. All isolates produced a cytotoxin in plasma active on polymorphonuclear (PMN) leucocytes. Cytotoxic proteins purified by FPLC exhibited a wide range of molecular weights. Various ELISA assays were developed for the detection of exotoxin A production. The incidence of toxin was similar in each group. Exotoxin A was rare in urines from infected patients, but was frequent in serum especially from chronic infections. Exotoxin A levels in sera of leukaemic patients was low. Antibody to exotoxin A was also low and anomalous results were obtained. Haemolytic activity was universal for all isolates and much of this was due to heat-stable glycolipid. A hypothetical model of interaction between exotoxin A production and cytotoxin is proposed in which the cytotoxin of P. aeruginosa cause lysis of leucocytes with the resultant release of leucocyte elastase enzyme which is highly active on exotoxin A. Neutropenic patients lack target cells for cytotoxin and thus exotoxin A activity is unchecked.
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Saulnier, Joëlle. "Activités enzymatiques de l'élastase de Pseudomonas aeruginosa." Lyon 1, 1989. http://www.theses.fr/1989LYO10107.
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Pseudomonas aeruginosa est une bacterie pathogene qui est responsable de nombreuses infections graves. Il a ete demontre que l'elastase secretee par la bacterie contribue a la pathogenicite de celle-ci mais son role exact n'a pas ete clairement elucide. C'est pourquoi nous avons entrepris d'etudier quelques unes de ses proprietes tant structurales qu'enzymatiques. Apres des rappels bibliographiques et la presentation des materiels et methodes utilises, nous avons expose nos resultats: 1) les vitesses initiales d'elastolyse d'elastines bovine et humaine par l'elastase ont ete mesurees par la methode conductimetrique. Il apparait que, quelle que soit l'elastine, l'affinite vis-a-vis de l'elastase de p. Aeruginosa est plus faible que pour les elastases pancreatique et leucocytaire mais l'activite est plus forte. 2) la stabilite thermique de l'elastase a ete etudiee lors d'experiences de denaturation et de desactivation thermique. L'elastase est stable (temperature de fusion principale d'environ 70#oc) et peu sensible a des effecteurs ioniques pour des concentrations de l'ordre du millimolaire. Par contre, meme a 30#oc, on peut mettre en evidence une desactivation, ce qui est en accord avec un modele a trois etats. 3) pour preciser la specificite de l'enzyme, nous avons synthetise et suivi l'hydrolyse de substrats synthetiques: furylacryloyl-tripeptides, oligomeres d'alanine et tetrapeptides du type ala-ala-x-ala. Il est apparu notamment que l'hydrolyse des oligomeres d'alanine est possible a partir de la tetraalanine, que l'augmentation du nombre d'alanine s'accompagne d'une augmentation de l'affinite et de l'activite. Enfin, dans le cas des tetrapeptides, c'est la leucine en position p1 qui entraine l'hydrolyse la plus efficace. La derniere partie a ete consacree a la mise au point du dosage conductimetrique de l'activite elastolytique dans des surnageants de cultures de p. A
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Vareechon, Chairut Charles. "Host-Pathogen Interaction in Pseudomonas aeruginosa Infection." Case Western Reserve University School of Graduate Studies / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=case1499427972888735.
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ANDRIEU, LAURENCE. "Caracterisation d'une alginate lyase de pseudomonas aeruginosa." Paris 11, 1995. http://www.theses.fr/1995PA112264.
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Pseudomonas aeruginosa est une bacterie pathogene opportuniste pouvant entrainer des infections graves, surtout au niveau des poumons des malades atteints de mucoviscidose. Sous l'effet de facteurs environnementaux, les bacteries vont produire des exopolysaccharides, les alginates, facteur important de la virulence de la bacterie. Les alginates sont des polymeres d'acides d-mannuronique et l-guluronique en liaison 1-4. Ils sont hydrolyses par un enzyme specifique, l'alginate lyase, qui catalyse une reaction de -elimination entre deux residus d-mannurosyl ou l-gulurosyl. Une activite alginolytique a ete mise en evidence chez deux souches mucoides de p. Aeruginosa. Cette activite est portee par une proteine homodimerique de 130 kda, intracellulaire en association avec la face interne de la membrane cytoplasmique. Le gene de structure de l'alginate lyase a ete clone a partir d'une banque d'adn genomique construite dans l'adn du phage lambda gt 11. Le gene algl, porte par un fragment de 4,4 kpb, est exprime chez e. Coli y 1090 en une proteine de 130 kda. Le sous-clonage de fragment de 4,4 kpb dans le gene cat du plasmide pbr 329 nous a permis de construire la carte de restriction du fragment, et de montrer que ce gene introduit chez e. Coli mc 1061, etait exprime, ce qui suggere que le gene a ete clone avec son propre promoteur. Les sous-clonages du fragment entier et de fragments de restriction, puis le clonage de fragments issus de l'hydrolyse controlee de l'insert de 4,4 kpb par la dnase i nous ont permis de determiner la totalite de la sequence nucleotidique. L'analyse de la sequence a revele la presence d'une phase ouverte de lecture de 2052 nucleotides, codant pour une proteine de poids moleculaire predit de 74 kda qui est en accord avec celui du monomere de l'alginate lyase determine par electrophorese sur gel sds, et qui doit correspondre au gene algl. De plus, la presence de sequences de fixation des ribosomes et de l'arn polymerase confirment que le gene algl possede son propre promoteur. Ces resultats montrent que l'alginate lyase mise en evidence est differente de celles caracterises chez d'autres souches de p. Aeruginosa
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Wilton, Alison Jane. "Iron-regulated surface antigens of Pseudomonas aeruginosa." Thesis, Aston University, 1989. http://publications.aston.ac.uk/12564/.
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Baretto, Bernadette J. "Studies on survival of Pseudomonas aeruginosa 6750." Thesis, Aston University, 1996. http://publications.aston.ac.uk/11046/.
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The growth of Pseudomonas aeruginosa 6750 as a biofilm was investigated using a novel system based on that of Gilbert et al (1989). The aim was to test the effect of controlled growth of the organism on antibiotic susceptibility and examine the survival of the organism as a biofilm. During the investigations it became clear that, because of the increasing growth of P.aeruginosa and production of exopolysaccharide, a growth rate controlled monolayer could not be achieved and so the method was not used further. The data, however, showed that there was an increase in the smooth colony type of the organism during growth. Investigations were focused on the survival of P.aeruginosa in batch and chemostat studies. Survival or percentage culturability, as measured by total and colony count ratio, was found to decrease both in extended batch culture and for chemostat cells with decreasing growth rate. Extended batch culture, however, did not exhibit further increases in resistance to ciprofloxacin and polymyxin B. Survival was also measured using other parameters namely the direct viable count, vital staining, effect of temperature downshift and measurement of lag. In batch culture, the most notable change was a decrease in cell size along the growth curve. This was accompanied by an increase in the cellular protein content. Protein per volume was calculated from the data which showed a marked increase in batch culture, which was not demonstrated for chemostat cells with decreasing growth rate. Outer membrane protein profiles were obtained for batch and chemostat cells. An LPS profile of batch culture cells was also demonstrated. In general, there was little difference in the outer membrane protein profiles of cells from early and late stationary phases.
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Noguès, Aurélie. "Résistance adaptative aux polymyxines chez Pseudomonas aeruginosa." Thesis, Besançon, 2015. http://www.theses.fr/2015BESA3006.
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La résistance aux polymyxines chez P. aeruginosa résulte en partie de la modification du lipide A par addition de 4-amino-aL-arabinose (L-Ara4N), due à l'expression de l'opéron arnBCADTEF-ugD (arn), activée par au moins 6 systèmes de régulation à deux composants (S2C). Nous avons mis en évidence que P. aeruginosa était capable de s'adapter de manière transitoire à la présence de fortes concentrations de polymyxines (8 x CMI) aussi bien in vitro que in vivo dans un modèle murin d'infection pulmonaire aiguë. La délétion de l'opéron arn chez la souche sauvage n'a pas modifié la capacité d'adaptation de P. aeruginosa. Afin d'identifier les gènes impliqués dans le processus adaptatif, le transcriptome global du mutant PAOl/z«;-Aar«-ATCS délété des principaux S2C (parRS, pmrAB, cprRS eiphoPQ) et de l'opéron arn a été réalisé en présence de différentes concentrations de colistine par RNA-Seq. Deux nouveaux mécanismes ont ainsi été identifiés. L'un repose sur l'expression de l'opéron mmsAB codant des enzymes du catabolisme des acides gras et l'autre fait intervenir le facteur sigma alternatif AlgU. Seule la délétion conjointe du gène algW participant à l'activation de AlgU et des opérons arn, mmsAB, pmrAB, parRS, phoPQ et cprRS a permis d'abolir complètement la résistance adaptative à la colistine. Par ailleurs, nous avons mis en évidence le rôle des vésicules de membrane externe (OMVs) dans la séquestration de l'antibiotique, dont la production semble régulée au moins par AlgU et le PQS. Ces travaux offrent des perspectives intéressantes pour l'identification de nouvelles cibles antibactériennes et pour l'amélioration de l'effet bactéricide des polymyxinesResistance to polymyxins in Pseudomonas aeruginosa involves the addition of 4-amino-L-arabinose (Ara4N) to LPS phosphates, thanks to an enzymatic modification due to the operon named arnBCADTEF-ugD (arn) whose expression is activated by at least 6 two component regulatory Systems (TCS). We demonstrated that P. aeruginosa was able to resist in a transient way to high concentrations of polymyxins (8x MIC) in vitro and in vivo in a mice lung infection model. Arn operon deletion in the wild type strain did not modify the ability to adapt to polymyxins. In order to identify gènes involved in adaptive resistance, we performed RNA-seq transcriptomes of quintuple mutant PAO\lux-Aaw-ATCS exposed to different concentrations of colistin or non exposed. Two new mechanisms were identified. The first one is based upon mmsAB operon encoding fatty acid catabolism enzymes and the second one is due to the sigma factor AlgU. Only the deletions of algW%enz involved in AlgU activation and arn, mmsAB, pmrAB, parRS, phoPQ and cprRS operons completely abolished the adaptive process. We also demonstrated the role of outer membrane vesicles in the sequestration of colistin whose production is regulated by AlgU and PQS. This study provides knoweldge essential for the design of novel strategies aimed at tackling the adaptive resistance to polymyxins
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Ferreira, Luciana Lobianco. "Estrutura clonal e multirresitência em Pseudomonas aeruginosa." reponame:Repositório Institucional da FIOCRUZ, 2005. https://www.arca.fiocruz.br/handle/icict/8542.
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Previous issue date: 2005Fundação Oswaldo Cruz. Instituto Nacional de Controle de Qualidade em Saúde. Rio de Janeiro, RJ, Brasil.
O presente trabalho teve como principal objetivo avaliar a multirresistência e os fatores envolvidos na resistência aos carbapenemas, em 187 cepas de Pseudomonas aeruginosa oriundas de hospitais no Rio de Janeiro {3) e Mato Grosso do Sul (1 ), e de dois laboratórios da rede de Saúde Pública (ES e BA). O mecanismo de resistência aos carbapenemas foi determinado através da pesquisa da produção de metalo-Beta-lactamase (Mbla) por meio de um teste fenotípico de difusão em duplo disco, utilizando EDTA, e a presença de dois genes responsáveis pela produção deste de tipo de Beta-lactamase, foi verificada através da PCR. No computo geral, 66.3 por cento das cepas mostraram-se resistentes a seis antimicrobianos ou mais. Resistência ao imipeném foi observada em 58,3 por cento dos isolados, e ao meropeném em 57.2 por cento, com 85.3 por cento de positividade para Mbla. A presença dos genes IMP-l e SPM foi detectada em 6.7 por cento e 34.5 por cento, respectivamente. Através da Eletroforese de Campo Pulsado foram identificados 93 genótipos distintos para as 187 cepas analisadas, apontando a prevalência de um perfil genômico (A), detectado em todas as regiões que participaram do estudo. Este se caracterizou por 96.5por cento de positividade para Mbla e 41.4 por cento para o gene SPM. O elevado percentual de multirresistência, e o achado de um genótipo multirresistente prevalente em diferentes regiões do Brasil, indicam a disseminação de genes de resistência o que representa uma grande preocupação principalmente quando isoladas no ambiente hospitalar, por poderem estar envolvidas com surtos e impedir o tratamento de primeira linha.
Nosocomial infections, many times are the main cause of morbity and mortality among hospitalized patients, configuring a serious problem by public health. In this context Pseudomonas aeruiginosa is a leading cause of this kind of infection. At Brazilian hospitals, Pseudomonas aeruginosa, ranked first among all nosocomial pathogens related to pneumonia in intensive care unit, emphasizing its wrapping up with pathologies. The situation is increasingly worst because Pseudomonas aeruginosa is becoming multirresistant, what is a limiting factor in the treatment. Among the antimicrobial agents that are used to treat infections caused by pathogens β-lactam-resistant, the carbapenems are very useful antimicrobial agent due to be stable to hydrolyzing by the most β-lactamases, including extended spectrum β-lactamases (ESBL). However, today is increasingly the number of P. aeruginosa resistant to this agent. Therefore, the aim of this study was evaluate the multirresistance and the factors that were involved at imipenem resistance. A total of 187 strains of Pseudomonas aeruginosa isolated at hospitals of Rio de Janeiro city (3) and Mato Grosso do Sul (1), and isolated in Public Helath laboratories situated at Espírito Santo and Bahia states. The mechanism to carbapenem-resistance was determined using a testing method for screening the production the metallo-β-lactamase (Mbla) using a convenient test using EDTA, and the detection of two genes producing metallo-β-lactamase was verified using PCR. Among all strains, 66.3% were resistant to 6 or more agents. Imipenem resistance was detected in 58.3%, and meropenem in 57.2%, where 85.3% strains was positive to the screening test to Mbla. IMP-1 was detected in 6.7% among the strains what were tested, and SPM-1 were detected in 34.5%. The Pulsed Field Gel Eletrophoresis (PFGE) identified 93 genotypes distinct among the 187 strains analyzed, and was observed the prevalence of one pattern (A), detected at all regions studied. This genotype was positive to the screening test in 96.5% and the SPM-1 gene was detected in 41.4% among the strains with this genotype. This high percentage of multirresistance and the presence of a predominant genotype common at the different regions of the Brazil, indicate the dissemination by resistance genes, representing a problem mainly when isolated at hospital, because the high possibility of an outbreak, impending the ideal treatment.