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1
Blankemeier, Andrew R. "Characterization of Pseudomonas fluorescens Biofilm." The Ohio State University, 2011. http://rave.ohiolink.edu/etdc/view?acc_num=osu1307731184.
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Hamel, Robert D. "Aluminum detoxification mechanisms in Pseudomonas fluorescens." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1997. http://www.collectionscanada.ca/obj/s4/f2/dsk2/tape16/PQDD_0025/MQ31433.pdf.
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Clements, Richard Steven. "The serological homology of Pseudomonas fluorescens proteases." Thesis, Queensland University of Technology, 1987. https://eprints.qut.edu.au/36720/1/36720_Clements_1987.pdf.
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This investigation evaluated the serological homology of
proteases from a number of psychotrophic bacteria
consisting largely of Pseudomonas_fluorescens strains.
Proteases from 4 reference strains of P.fluorescens were
purified and rabbit antiserum generated against each.
Preliminary investigations revealed the serological
heterology of protease OM82 to proteases N73A, M143A and
OMl 86.
A total of 54, presumably P.fluorescens strains, were
grown and semi-pure protease preparations obtained from
each.
The immunological cross reactivity of 54 semi-pure
protease preparations with each of the 4 antisera was
evaluated using Ouchterlony Double Diffusion (ODDT),
Single Radial Immunodiffusion (SRID) and an Inhibition
Enzyme-Linked Immunosorbent Assay (IELISA).
The ODDT and IELISA were qualitative tests however,
densitometric analysis of SRID precipitin rings enabled
cross reaction quantitation in this assay.
Each protease was placed into one of 3 enzymo-serogroups
according to. the results from each immunological assay.
Proteases which did not cross react with any antisera
could not be classified.
A recent taxonomic study (O'Connor ~t _ gl., 1986)
indicated that
investigation
~.fluorescens.
produced by non
several of the enzymes used in this
were not produced by strains of
It was generally found that proteases
P.fluorescens species did not cross react
with reference P.fluorescens proteases.
Further development of the IELISA increased sensitivity
thus, enabling the detection of low nanogram levels of
specific proteases in ultra high temperature treated
milk. A number of monoclonal antibodies were produced and used
to determine whether a common antigenic determinant
existed amongst these proteolytic enzymes . Evi dence
presented in this study suggests the existence of such an
epitope however, the distribution of this determinant
amongst the secreted proteases of P . flyore~.Qens is not
known .
4
Wang, Chien-Sao. "Cell-Free Recovery and Isotopic Identification of Cyanide Degrading Enzymes from Pseudomonas Fluorescens." Thesis, University of North Texas, 1995. https://digital.library.unt.edu/ark:/67531/metadc278363/.
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Cell-free extracts from Pseudomonas fluorescens NCIMB 11764 catalyzed the degradation of cyanide into products that included C02, formic acid, formamide and ammonia. Cyanide-degrading activity was localized to cytosolic cell fractions and was observed at substrate concentrations as high as 100 mM. Two cyanide degrading activities were identified by: (i) the determination of reaction products stoichiometries, (ii) requirements for NADH and oxygen, and (iii) kinetic analysis. The first activity produced CO2 and NH3 as reaction products, was dependent on oxygen and NADH for activity, and displayed an apparent Km for cyanide of 1.2 mM. The second activity generated formic acid (and NH3) pfus formamide as reaction products, was oxygen independent, and had an apparent Km of 12 mM for cyanide. The first enzymatic activity was identified as cyanide oxygenase whereas the second activity consists of two enzymes, a cyanide nitrilase (dihydratase) and putative cyanide hydratase. In addition to these enzymes, cyanide-grown cells were also induced for formate dehydrogenase (FDH), providing a means of recycling NADH utilized by cyanide oxygenase.
5
Levasseur, Rémi. "Aluminum citrate transport and metabolism in Pseudomonas fluorescens." Thesis, National Library of Canada = Bibliothèque nationale du Canada, 1999. http://www.collectionscanada.ca/obj/s4/f2/dsk1/tape7/PQDD_0018/MQ46489.pdf.
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Wan, Dagang Wan Rosmiza Zana. "Understanding adhesion of Pseudomonas fluorescens on household surfaces." Thesis, University of Birmingham, 2012. http://etheses.bham.ac.uk//id/eprint/3821/.
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In this study, three different methods have been used to investigate the bacterial interaction with the substratum, i.e. atomic force microscopy (AFM), spinning disc and micromanipulation. Pseudomonas fluorescens NCIMB 9046 was chosen as a model microorganism to study the cell-substrate adhesion. By having three different colloidal particles: stainless steel (Grade 304), glass and cellulose, the force measurements were performed in growth medium and ambient air using AFM. The results demonstrated that the adhesive forces were influenced by the surface hydrophobicity, electrostatic, van der Waals and steric interactions. In ambient air, the capillary force played an important role. The effect of shear forces on the bacterial adhesion was further examined. By using an apparatus of spinning disc, the cell removal was strongly influenced by the spinning time, angular velocity and surface hydrophobicity. Finally, the adhesive and cohesive strengths of biofilms were examined via a micromanipulation technique. Results indicate that with pH7 and low initial glucose concentration (0.25% (w/v)) the biofilm adhesion was the greatest among the conditions investigated. The cohesive strength of biofilm was found to depend on on the distance between the force probe and the substrate surface.
7
Macioszek, Malgorzata. "Biosynthesis of mupirocin by Pseudomonas fluorescens NCIMB 10586." Thesis, University of Birmingham, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.510237.
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Mupirocin, a polyketide antibiotic active against Gram-positive bacteria is used clinically for treatment of bacterial skin infections, to clear Stapylococcus aureus ftom nasal passages and as a surgical scrub to inhibit bacterial growth, particularly that of MRSA. Mupirocin is synthesised by polyketide synthases (PKS) in a series of reactions involving many enzymes encoded by genes from the mupirocin cluster. The mupirocin cluster consists of six larger ORFs (mmpA-F. ) encoding multifunctional proteins, and twenty nine individual genes (mupA-X and macpA-E) all of which have been shown to be required for normal mupirocin biosynthesis and presumably create a biosynthetic assembly line. Sub-groups of mutants produce identical novel metabolites implying the presence of multi-protein complexes. To determine the interactions between proteins of the Mup assembly line Bacterial and Yeast Two Hybrid Systems were used but no evidence has been provided that tested proteins are partners. Although no interaction has been revealed, complementation studies suggest that MupE protein requires coexpression of MupD protein to be functional. Furthermore, inactivation of MupD by amino acid substitution suggests that MupD is not essential on its own for any step in mupirocin biosynthesis except for the proper function of MupE protein. BPLC analysis of wild type P. fluorescens overexpressing mupOlmacpElUIVICIF in trans showed that this did not abolish PA-B production as had been hypothesised but did increase production of total antibiotic (PA-A and PA-B) up to more than two-fold indicating a need to modify our model for the production of PA-B. Results using fluorescence microscopy demonstrate that small Mup proteins are not localized within the bacterial cell and that they are spread evenly in the cells of P. fluorescens.
8
Frey-Klett, Pascale. "Ecologie d'un pseudomonas fluorescens auxiliaire de la mycorhization." Paris 11, 1996. http://www.theses.fr/1996PA112480.
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La souche de bacterie auxiliaire de la mycorhization pseudomonas fluorescens bbc6, isolee d'un carpophore de laccaria laccata, stimule l'etablissement de la symbiose entre le douglas et ce champignon ectomycorhizien. Dans le but de comprendre son mode d'action mais aussi pour integrer de facon raisonnee son inoculation aux programmes de mycorhization controlee du douglas, nous avons etudie l'ecologie de la souche bbc6 en serre et en pepiniere. Nous avons compare les caracteristiques phenotypiques et genotypiques de bbc6 a celles de 300 souches de pseudomonas fluorescents isolees du sol nu, de la mycorhizosphere et des mycorhizes de douglas-l. Laccata dans un essai en pepiniere. Bbc6 appartient au biovar i des p. Fluorescens et partage des caracteristiques phenotypiques communes a toutes les souches de p. Fluorescens biovar i isolees de la mycorhizosphere et des mycorhizes. Elle est capable d'utiliser le trehalose, sucre majoritairement accumule dans le mycelium de l. Laccata in vitro. Nous avons d'autre part etudie l'ecologie de bbc6 grace a l'utilisation d'un mutant spontane resistant a la rifampicine. La population bacterienne decroit dans le sol et la rhizosphere des semis de douglas apres l'inoculation. La bacterie n'est donc ni tellurique, ni rhizospherique. Elle n'est pas non plus preferentiellement associee aux mycorhizes ni aux carpophores de l. Laccata. Pourtant, la bacterie survit mieux en presence de l. Laccata. La bacterie pourrait donc etre associee au mycelium du champignon dans le sol et profiter des exsudats fongiques, comme le trehalose par exemple. L'effet auxiliaire de la bacterie se manifeste trois a six semaines apres la formation des premieres mycorhizes, lorsque la densite bacterienne a deja chute d'au moins 1000 fois. Il se manifeste egalement pour des doses d'inoculum bacterien aussi faibles que 10 ufc cm#-#3 de sol et permet de diminuer la dose d'inoculum fongique utilisee sans diminuer l'indice de mycorhization des plants
9
Koza, Anna. "Adaptation and niche construction by Pseudomonas fluorescens SBW25." Thesis, Abertay University, 2011. https://rke.abertay.ac.uk/en/studentTheses/7888a5ac-f562-4518-8124-36d2e394994d.
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The mechanisms underlying adaptive radiation or evolution have been extensively investigated using experimental bacterial populations in liquid cultures, referred to as microcosms. Evolving populations of Pseudomonas fluorescens SBW25 in static microcosms reproducibly lead to the emergence of the Wrinkly Spreader (WS) genotype. These produce a cellulose-matrix-based biofilm to colonise the airliquid (A-L) interface with significant fitness advantage over non-biofilm-forming competitors. In this work, the first SBW25 colonists in static microcosms were shown to establish O2 gradients, thereby modifying the original environment to establish two new niches corresponding to a high-O2 region at the A-L interface and a low-O2 region lower down the liquid column. Greater O2 availability at the A-L interface was found to provide the selective pressure driving the emergence of the WS and underlay the fitness advantage WS have over non-biofilm-forming strains in this environment. A second biofilm-forming mutant of SBW25, known as the CBFS (Complementary Biofilm Forming Strain), had been previously characterised. Here, wild-type SBW25 was shown to produce a third biofilm-type induced non-specifically by exogenous Fe, and referred to as the viscous mass (VM)-class biofilm. The CBFS, VM and WS biofilm-types could be differentiated by in situ measurements of biofilm attachment and strength, rheometry of biofilm samples, and strain characteristics. However, despite these differences, each provided similar levels of fitness in static microcosms subjected to increasing levels of physical disturbance. This suggests that each of the biofilm-types might arise independently in evolving SBW25 populations as the result of convergent evolution, in which the key ecological constraints are O2 availability and physical disturbance. However, invasion-from-rare experiments indicated that the WS was ecologically more successful, perhaps explaining why only WS-like genotypes have been isolated from evolving SBW25 populations in static microcosms. Other pseudomonads had previously been shown to produce cellulose-based A-L interface biofilms in static microcosms. Here, a survey of New Zealand brown blotch-causing pseudomonads (BCP) recovered from white mushrooms (Agaricus bisporus) identified similar biofilm-producing, cellulose-expressing isolates. The ability to express cellulose by key BCP isolates was shown to be a fitness advantage in static microcosms, as well as on A. bisporus mushroom caps themselves. Cellulose-expression was also found to be of fitness advantage under water-limiting conditions, suggesting that cellulose may provide some resistance to water-stress in the natural environment. The research undertaken for this Thesis comprise several significant advances in our understanding of the adaptive radiation of P. fluorescens SBW25 and the emergence of A-L interface biofilms in static microcosms. This work has identified O2 gradients and physical disturbance as the dominant environmental factors that guide the convergent evolution of biofilms in this environment. A consequence of convergent evolution in static microcosms is a variety of physically-different biofilms, all of which provide a fitness advantage over non-biofilm-forming competitors. It is also enlightening to realise that a common structural component of biofilms, cellulose, may also be used in another role to provide a fitness advantage in an ecologically more relevant and natural environment.
10
Kulkarni, N. "Studies on lipase enzyme from pseudomonas fluorescens NS2W." Thesis(Ph.D.), CSIR-National Chemical Laboratory, Pune, 2002. http://dspace.ncl.res.in:8080/xmlui/handle/20.500.12252/2333.
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Delafuente, Leonardo. "Characterization of the ecological and physiological basis of superior rhizosphere colonization by 2,4-diacetylphloroglucinol-producing fluorescent Pseudomonas genotypes." Online access for everyone, 2005. http://www.dissertations.wsu.edu/Dissertations/Fall2005/l%5Fdelafuente%5F1082405.pdf.
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Uría, Fernández Diana. "Strukturaufklärung der Siderophore und massenspektrometrische Untersuchung eines ihrer Rezeptorproteine von Pseudomonas fluorescens G173." [S.l.] : [s.n.], 2002. http://deposit.ddb.de/cgi-bin/dokserv?idn=964735342.
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Janzen, Elena. "Die Benzaldehydlyase aus Pseudomonas fluoreszens biochemische Charakterisierung und die Untersuchung von Struktur-Funktionsbeziehungen /." [S.l. : s.n.], 2002. http://deposit.ddb.de/cgi-bin/dokserv?idn=967208629.
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14
Jackson, Benjamin. "Molecular genetics of #alpha# pinene metabolism in Pseudomonas fluorescens." Thesis, Imperial College London, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.343598.
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15
Scott, Robert W. "Synthetic strategies to metabolites from mutants of pseudomonas fluorescens." Thesis, University of Bristol, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.558095.
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Mupirocins F and H are metabolites isolated from cultures of mutant strains of Pseudomonas f1uorescens. Synthetic routes have been developed to these compounds to confirm their structures Investigations into the synthesis of 22 led to two novel rearrangement products 48 and 55 and their structures were confirmed through extensive spectroscopic studies. Methyl mupirocin F was prepared in two steps from pseudomonic acid A via methylation with TMS- diazomethane to give methyl pseudomonate 3 followed by a selective TEMPO oxidation to yield methyl mupirocin F 22. The total synthesis of mupirocin H 73 has been achieved using a flexible synthetic strategy. Lactone 85 was prepared from bicycloheptanone 88 which on alkylation with 86 gave 186 with complete stereocontrol and all six asymmetric centres intact. Reductive cleavage of lactone 186 via the intermediate dithiane 199 installed the required methyl group and oxidation gave 206. Removal of the protecting groups gave mupirocin H 73.
16
Griffiths, M. B. "Genetic analysis of ecgonine degradation by Pseudomonas fluorescens (MBER)." Thesis, University of Cambridge, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.599726.
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This study details a genetic investigation of ecgonine degradation by P. fluorescens MBER. Transposon (Tn5) insertional inactivation mutagenesis was performed to create mutants blocked in the ecgonine degradative pathway. By this method, 87 such mutants were isolated and analysed to identify where in the genome the interruption had occurred. At the whole cell and cell-free extract level, no intermediary metabolites were detected in these mutants by either TLC or GC/MS analysis. The genetic location of Tn5 in the DNA was therefore investigated to obtain sequence data around the insertion site which might identify the gene into which insertion occurred. This genetic analysis revealed that the transposon had inserted in an identical fragment in all 87 mutants, indicating a hyper-insertion region in the DNA. Sequence data flanking the insertion site did not exhibit homology to any known sequences in world-wide databases; the identity of the interrupted gene currently remains unknown. Co-isolated with the unidentified sequence, however, was DNA exhibiting high homology to a phenol hydroxylase, and a benzene monooxygenase. Initial enzyme induction studies revealed that the meta-cleavage pathway associated with catechol metabolism in P. fluorescens MBER was induced by growth on ecgonine as the sole source of carbon. Putative evidence obtained during the course of this study infers that aromatic degradative enzymes may be involved in the later stages of ecgonine degradation. The results obtained are also discussed in the light of subsequent work, which discovered the unidentified DNA interrupted by Tn5 insertion resulting in loss of ecgonine degrading ability was not co-located with the putative benzene monooxygenase and phenol hydroxylase.
17
Reid, H. D. "An investigation of Pseudomonas fluorescens in the milk environment." Thesis, University of Canterbury. Microbiology, 1997. http://hdl.handle.net/10092/6873.
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The main aim of this study was to determine the percentage of Pseudomonas fluorescens amongst fluorescent pseudomonads in the milk environment, and to distinguish these isolates from each other. A secondary aim was to investigate whether these isolates were clonal or non-clonal in origin, and to study whether end product contamination had an endemic factory source.
In this study 230 fluorescent bacteria were isolated from a South Island dairy company over a six month period. These bacteria were isolated from raw milk and associated environments, separated milk and associated environments, pasteurised milk environments and from final product sampling. These isolates were biotyped using the API-20NE biochemical test system. Approximately 70% of these isolates were identified as Ps. fluorescens, and the majority of these grouped into the biotypes 0157555 and 0357555. Strains of Ps. fluorescens grouped into these biotypes were the only proteolytic enzyme producers isolated from final product isolates.
To investigate whether end product contamination had an endemic factory source, 97 strains of the biotypes 0157555 and 0357555 were characterised using the technique of multilocus enzyme electrophoresis. These strains clustered into 26 distinct electromorphs, with the majority of isolates falling into one cluster. Isolates in this cluster originated from raw milk and associated environments, separated milk and associated environments, the pasteurised milk environment and the final product. With the exception of month four there is a constant presence of the major cluster strains in the milk environment. This cluster can be divided further as both of the biotypes studied (0157555 and 0357555) were present in this group. Isolates in this major cluster could be clonal in origin and represent a serious contaminating organism able to occupy many niches in the milk processing environment over a considerable time period.
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McCloskey, Nicholas. "Transcriptional Effects of Adaptive Synonymous Mutations in Pseudomonas fluorescens." Thesis, Université d'Ottawa / University of Ottawa, 2018. http://hdl.handle.net/10393/37903.
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Synonymous mutations have traditionally been thought to have no significant effect on fitness. However, a growing body of recent research has shown that this is not always the case. In an experimentally evolved population of Pseudomonas fluorescens grown in minimal glucose media, synonymous mutations arose in a glucose transport gene that resulted in beneficial fitness effects comparable to those of non-synonymous mutations. We found that the increase in fitness was a direct result of increased gene expression; however, the precise mechanism was unclear. Synonymous mutations have been shown to affect gene expression on transcriptional and translational levels through changes in mRNA secondary structure and codon usage.
Our study investigates the underlying mechanisms in which these evolved synonymous mutations lead to increased gene expression. In addition to the evolved mutations, we have a library of 42 strains with single synonymous mutations within the glucose transport gene and found a positive correlation between fitness and gene expression. To determine whether these mutations affect transcript levels, translational efficiency or a combination of both, we systematically incorporated transcriptional and translational fusions of a yellow fluorescent protein within the glucose transport operon. We found that the evolved mutations predominantly act on the level of transcription and have strong polar downstream effects. Additionally, through manipulation of the local genetic sequence, we investigated the specific molecular requirements necessary for the increased expression. We found that for one of our evolved synonymous mutants, mRNA secondary structure does not play an essential role, but we speculate that the mutation may strengthen a weak internal promoter sequence to confer its increased expression. Our study provides evidence of the adaptive mechanisms of beneficial synonymous mutations in an experimentally evolved setting.
19
Nagappan, Olagappan. "Mechanisms of Cyanide Assimilation in Pseudomonas fluorescens NCIMB 11764." Thesis, University of North Texas, 1992. https://digital.library.unt.edu/ark:/67531/metadc278533/.
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Pseudomonas fluorescens NCIMB 11764 was capable of utilizing cyanide as a sole nitrogen source for growth. Cyanate (OCN") and S-cyanoalanine could also serve as nitrogenous substrates, but do not appear to play a role as intermediates in cyanide metabolism. Growth of this strain on cyanate as the sole nitrogen source led to the induction of an enzyme characterized as a cyanase (EC 3.5.5.3) based on its stoichiometric conversion of cyanate to ammonia, and dependence on bicarbonate for maximal activity. However, since cyanase activity was not elevated in cyanide-grown cells it was concluded that it serves no role in cyanide metabolism. Related studies aimed at examining a possible role for S-cyanoalanine as a cyanide-assimilation intermediate showed that while this compound also serves as a nitrogen source, it also is not important in cyanide metabolism. Studies focused on the utilization of free cyanide as a growth substrate led to the development of a fed-batch cultivation procedure greatly facilitating further experimentation aimed at the identification of cyanide metabolites. In addition to CO_2 and NH_3 as described earlier, two additional metabolites including formamide and formate were detected by using nC-NMR, HPLC, radioisotrapping methods and other analytical means. The formation of metabolites was shown to be induced after growth on cyanide with the relative product yields dependent on the availability of oxygen. These findings support earlier work in which an oxygen-dependent mechanism was proposed for the formation of C02 and NH3. However, at least two additional oxygen-independent pathways of cyanide conversion can be elaborated by this organism. One of these involves conversion to formate and ammonia while the other leads to the formation of formamide, which is not further degraded. Thus, growth on cyanide appears to occur by several mechanisms of chemical transformation presumably serving both detoxification and nutritional roles. Since two of these mechanisms generate ammonia, which is readily assimilated, growth is presumed to proceed via ammonia as a provisionary nitrogenous substrate.
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Lin, Hao. "PREDICTION OF GROWTH OF PSEUDOMONAS FLUORESCENS UNDER TEMPERATURE FLUCTUATION." The Ohio State University, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=osu1429840331.
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Senatore, Marcela Andrea Duran Haun 1974. "Avaliação da eficiencia de um esterilizador a plasma na inativação de Pseudomonas fluorescens." [s.n.], 2004. http://repositorio.unicamp.br/jspui/handle/REPOSIP/254838.
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Orientador: Marcelo CristianiniDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia de Alimentos
Made available in DSpace on 2018-08-03T20:56:17Z (GMT). No. of bitstreams: 1 Senatore_MarcelaAndreaDuranHaun_M.pdf: 684290 bytes, checksum: 522970fe79261c7d2890c0a19a39b587 (MD5) Previous issue date: 2004
Resumo: Um dos problemas que enfrenta a indústria de laticínios é a recontaminação do leite decorrente da formação de biofilmes em tanques de armazenamento e trocadores de calor. A tecnologia de esterilização de materiais por gás plasma tem sido utilizada com sucesso na esterilização de instrumentos cirúrgicos, oftálmicos e dentários. Os objetivos deste trabalho foram avaliar a eficiência de um sistema de esterilização a gás plasma sob células de Pseudomonas fluorescens aderidas em placas de aço inoxidável utilizando o leite como substrato. Foram avaliadas as variáveis tempo de pré plasma, tempo de exposição ao plasma e potência do mesmo na remoção do biofilme. As placas de aço inoxidável com a bactéria aderida foram submetidas a um tratamento com gás plasma, que foi formado a partir de um produto comercial composto por ácido peracético, peróxido de hidrogênio e ácido acético. Dois sistemas modelo foram utilizados para simular a formação de biofilmes de Pseudomonas fluorescens, um dinâmico e outro estático, simulando trocadores de calor e tanques de armazenamento, respectivamente. Através do modelo estático foi possível obter contagens acima de 105 UFC por placa de aço inoxidável, mesmo após três ciclos de lavagem das placas em água estéril sob agitação. Foi observado um efeito positivo na inativação de P. fluorescens em função do tempo de pré-plasma do sanificante. Exposições acima de 7 minutos foram capazes de produzir reduções superiores a dois ciclos logarítmicos na inativação do microrganismo. Através de um planejamento experimental fatorial 23 foi demosntrado que as variáveis tempo de pré-plasma, tempo de exposição ao plasma e potência do plasma apresentaram efeitos positivos na inativação de Pseudomonas fluorescens. O tempo de exposição (min.) apresentou o maior efeito na destruição da bactéria; mas sendo um pouco superior à potência do plasma (w)
Abstract: One of the problems that food industry is facing is the milk recontamination through biofims formation on storage tanks and heat exchanger. The technology of sterilization for materials through gas plasma has been used with succesfull for cirurgycal, ophathalmic and dentistry equipment. The goals of this work was to evaluate the efficiency of a gas plasma sterilization system on Pseudomonas fluorescens cells adhered on stainless steel plates using milk as substrate. Time of pre plasma, plasma exposition and potency (Watts) were evaluated as independent variables on cell destruction. The stainless steel plates were submitted to gas plasma treatment originated from a commercial product composed of peracetic acid, hydrogen peroxide and acetic acid. Two model systems were used to simulate a biofilm formation of Pseudomonas fluorescens, one dynamic and one static, in order to simulate heat exchangers and storage tanks, respectively. Through static model it was possible to get counts over 105 UFC/plate after washing three times using sterile water under stirring conditions. It was observed a positive effect on P. fluorescens inactivation by pre-plasma in a time dependent way. Expositions over seven minutes were capable to produce reductions higher than two logarithmic cycles on microorganism inactivation. A 23 factorial design indicated that the pre-plasma time, time exposition and potency showed positive effects on Pseudomonas fluorescens inactivation. Time exposition (min) was the most effective variable on bacteria destruction, being a little higher than plasma potency (w)
Mestrado
Mestre em Tecnologia de Alimentos
22
Barrett, Rowan Douglas Hilton. "Experimental evolution of Pseudomonas fluorescens in simple and complex environments." Thesis, McGill University, 2005. http://digitool.Library.McGill.CA:80/R/?func=dbin-jump-full&object_id=97902.
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Determining the factors responsible for the origin and maintenance of diversity remains a difficult problem in evolutionary biology. There is extensive theoretical work which suggests that environmental heterogeneity plays a major role. This theory argues that diversification is ultimately due to divergent natural selection for alternative resources. In this thesis I investigate adaptation and the evolution of diversity in experimental populations of the asexual bacterium Pseudomonas fluorescens. In all experiments I introduce clonal isolates of Pseudomonas to a novel environment and allow evolution to occur through the substitution of random mutations. Adaptation can then be quantified by comparing evolved genotypes to the ancestor. These experiments show that when Pseudomonas is selected in a complex environment containing several resources, sympatric genotypes adapt to use different resources, leading to the evolution of genetically diverse populations. In environments containing just a single resource, most genotypes adapt to use the same resource and no such diversity is observed. Adaptation in the experimental populations is caused by the fixation of beneficial mutations of intermediate fitness effect. My results highlight the value of microbial model systems for answering evolutionary questions and provide strong evidence for the role of ecological factors in the origin of diversity.
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Turnbull, Gillian Anne. "The role of motility in Pseudomonas fluorescens and Pseudomonas putida in soil-plant-microbe interactions." Thesis, University of Liverpool, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.367105.
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Kondakova, Tatiana. "Pseudomonas adaptation to stress factors : role of membrane lipids and Pseudomonas fluorescens response to NO2." Rouen, 2015. http://www.theses.fr/2015ROUES016.
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La large distribution des Pseudomonas fluorescens est liée à leur grande adaptabilité aux facteurs de stress tels que les variations environnementales. Ce travail avait pour objet la réponse spécifique au milieu aéroporté de P. Fluorescens, comme l’aéroportée MFAF76a et la clinique MFN1032 comme standard. La technique récente HPTLC-MALDI-TOF MSI a permis de caractériser les divers glycérophospholipides (GP) des deux souches. En phase stationnaire de croissance, un GP inconnu (UGP - unknown GP) a été isolé et semble intervenir dans l’adaptation à la température de la souche clinique MFN1032. Quant au stress NO2 gaz, les deux souches ont été exposées aux concentrations: 0. 1, 5 et 45 ppm. Leurs phénotypes ont été confrontés à l’expression de quelques gènes ciblés. Pour les valeurs standard 0,5 and 5 ppm en NO2, aucun paramètre n’est modifié. Par contre, une réponse bactérienne est constatée suite à l’exposition à 45 ppm de NO2. Cette exposition semble impacter la production d’UGP. De plus hmp et mexEF-oprN, codant respectivement pour la flavohémoglobine et pour la pompe à efflux RND, se trouvent surexprimés, corroborant l’évolution de la résistance bactérienne aux antibiotiques. Contrairement au NO, aucune altération de la biomasse de biofilm n’est observée pour le NO2, qui favorise cependant l’augmentation de son épaisseur chez MFAF76a, mais aussi l’inhibition du swarming et la diminution du swimming, avec un taux de c-di-GMP stable. Ce faisceau de résultats offre, pour la première fois, la réponse bactérienne et le rôle des GP lors de stress comme NO2 ou à la température, autre modification environnementaleThe high distribution of Pseudomonas fluorescens group is linked to its ability to adapt to stress factors. This work goaled the response of an airborne P. Fluorescens MFAF76a, and its clinical standard P. Fluorescens MFN1032 to environmental changes in order to refine the specific adaptation of airborne bacteria. First the HPTLC-MALDI TOF MSI tool defined glycerophospholipid (GP) composition of both strains. In stationary growth phase, an unknown GP, in short UGP, was found and seemed to be involved in temperature adaptation for the clinical strain. After exposure to 0. 1, 5 and 45 ppm concentrations, the bacterial response to NO2 was defined through motility, biofilm formation, antibiotic resistance and expression of several chosen target genes. While no change in parameters was seen in bacteria exposed to 0. 1 and 5 ppm of NO2, several alterations were occurred with a bacterial exposure to 45 ppm. NO2 seemed to bias the UGP production, reduced P. Fluorescens swim and decreased swarm only for MFN1032 strain. Biofilm formed by NO2-treated MFAF76a showed increased maximum thickness, with no change in c-di-GMP intracellular level. Expression of the hmp-homologue gene involved in NO detoxification was upregulated in response to NO2, suggesting a possible common pathway between NO and NO2 detoxification. Finally, NO2 was found to increase bacterial resistance to ciprofloxacin and chloramphenicol. Thus the resistance nodulation cell division (RND) MexEF-OprN efflux pump encoding genes were highly upregulated in both strains. Together these findings implement the first model of bacterial response to NO2 toxicity and the role(s) of GP in bacterial adaptation to environmental changes
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Le, Clanche Jean-François. "La petite agriciculture : survivance du passé ou agriculture en devenir ? : Une approche bioéconomique." Rennes, Agrocampus Ouest, 2013. http://www.theses.fr/2013NSARE035.
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Petites exploitations, petits exploitants: ces mots éveillent en nous l’image d’une paysannerie aujourd’hui disparue, d’une agriculture condamnée par la modernité. D’autres esprits s’enflamment et y voient, à contrario, une issue probable pour dépasser les failles actuelles du modèle de développement productiviste. Si le nombre de petites exploitations a chuté vertigineusement depuis plus d’un siècle, elles représentent encore entre 20 et 25% du nombre total d’exploitations. Les économistes prévoyaient leur disparition. Leur prévision s’est même faite certitude jusqu’à s’imposer comme étant l’image du réel. Force est de constater que leur prédiction ne s’est pas réalisée. Contre toute attente, la petite agriculture résiste aux mécanismes qui contribuent à sa disparition. Pour mieux comprendre ces divergences et certains clivages idéologiques, cette thèse s’est centrée sur une analyse épistémologique de la diversité des approches socio-économiques traitant de la petite agriculture en s’ancrant notamment dans le courant de la bioéconomie. Elle retient l’hypothèse tchayanovienne qu’on ne peut pas étudier une exploitation agricole en l’assimilant à une entreprise et en mobilisant l’orthodoxie de l’analyse néo-classique. Diverses, explorant parfois des voies s’éloignant de la modernité, les petites exploitations peuvent être pérennes et s’inscrire dans une dynamique de recherche d’une plus grande durabilité. Certaines sont innovantes et pilotées par des agriculteurs qui mériteraient d’être qualifiés d’« entrepreneur schumpéterien ». On peut alors s’interroger sur la nature des politiques publiques qui, depuis plus d’un demi-siècle, s’obstinent à privilégier une forme d’agriculture et un type de structure, « l’exploitation familiale à deux U. T. H ». Si la création d’emplois est un objectif suivi, si l’on mobilise des valeurs de justice et d’équité, ce sont les fondamentaux de cette politique qu’il faut revisiter au niveau européen, national et territorial. La dernière partie de ce travail fait des propositions allant dans ce sensSmall farms: these words awaken in us the representation of a farming community today disappeared which the modernity of the post-war years would have condemned. Other spirits ignite and see, in contrario, an issue there to exceed the weaknesses of the model of productivist development in agriculture. If the number of small farms fell vertiginously for a century, they still represent between 20 and 25 % of the total number. The economists planned their disappearance. Their forecast was even made certainty until stand out as being the image of the reality. One has to note that their prediction did not come true. Contrary to all expectations, small farms resist the mechanisms which contribute in their disparition. To understand better these differences and certain ideological splits (cleavages), this thesis centered on an epistemological analysis of the diversity of the socioeconomic traitants approaches of the small agriculture (farming) by anchoring in particular in the course of the bioeconomy. It retains the hypothesis of A. Tchayanov that we cannot study a farm by assimilating it to a firm (enterprise) and by mobilizing the orthodoxy of the neo-classic analysis. Diverse, sometimes investigating ways going away from the modernity, the small farms can be long-lasting and join a dynamics of research for more durability. Some are innovative and piloted by farmers who would deserve to be qualified as “schumpétérien entrepreneur”. We can then wonder about the nature of the public policies which, for more than half a century, persist in favoring a shape of agriculture and a type of structure, “the family exploitation with two U. T. H”. If the job creation is a followed objective, if we mobilize values of justice and equity, it is the fundamental of this policy which it is necessary to revisit at the European, national and territorial level. The last part of this work makes proposals going to this sense (direction)
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Lou, Chun-hin. "Multilocus sequence typing of pseudomonas fluorescens isolates from investigation of a case of transfusion-associated sepsis." Click to view the E-thesis via HKUTO, 2009. http://sunzi.lib.hku.hk/hkuto/record/B42925320.
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Leneveu-Jenvrin, Charlène. "Virulence et adaptation à l'environnement de souches de Pseudomonas fluorescens et Pseudomonas mosselii ; contribution à l'étude des conditions d'expression et des fonctions de la protéine TSPO chez P. Fluorescens." Rouen, 2014. http://www.theses.fr/2014ROUES033.
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Ce mémoire est dédié d'une part à l'étude de la virulence de Pseudomonas mosselii, et d'autre part à l'étude du rôle de la protéine TSPO chez Pseudomonas fluorescens Pf0-1. P. Mosselii est une bactérie isolée du milieu hospitalier qui a été caractérisée en 2002 mais dont le pouvoir pathogène reste inconnu à ce jour. Nous avons pu observer que P. Mosselii ATCC BAA-99 et P. Mosselii MFY161 sont cytotoxiques, peu invasives mais capables d'altérer la perméabilité épithéliale des cellules Caco-2/TC7 et d'endommager le cytosquelette d'actine. Leur comportement ressemble à celui de certaines souches cliniques de P. Fluorescens. En raison de leur implication éventuelle en tant que vecteur de transfert de gènes de résistance aux beta-lactamines, ces bactéries mériteraient une surveillance accrue. Le pouvoir cytotoxique de P. Fluorescens Pf0-1 a également été testé et il s'est révélé faible dans nos conditions expérimentales. Par contre, cette bactérie nous a intéressé par le fait qu'elle possède une protéine translocatrice (TSPO), largement conservée au cours de l'évolution (Eukarya, Archae et Bacteria), mais dont le rôle chez cette bactérie était inconnu. La suite de nos travaux a donc consisté en l'étude de l'expression de cette protéine et de sa (ses) fonction (s) potentielle(s). Nous avons pu montrer la présence d'une histidine kinase hybride (Pfl01_2810) en amont du gène tspo avec laquelle tspo est co-transcrit. Cet opéron est exprimé de façon transitoire pendant la croissance bactérienne en milieu LB à 28°C. L'activité transcriptionnelle varie avec la température, elle est nulle à 18°C et très largement augmentée à 32°C, suggérant une régulation par le facteur sigma RpoH, généralement activé en réponse au stress thermique. En cas d'hypersmolarité (NaCl, Sucrose), ou en présence de D-cyclosérine, nous avons pu observer que l'activité transcriptionnelle est très largement augmentée, engendrant un stress pariétal et une régulation par le facteur sigma AlgU via RpoH. L'utilisation de lignads de la TSPO mitochondriale tel que le cholestérol, qui est impliqué dans la fluidité membranaire, engendre également une augmentation de la transcription de tspo, ainsi que d'autres ligands Eucaryotes tels que le PK11195 ou la protoporphyrine IX suggérant une conservation fonctionnelle entre la protéine TSPO bactérienne de P. Fluorescens Pf0-1 et son orthologue EucaryoteThis work aims to study the virulence of Pseudomonas mosselii and the role of TSPO protein in Pseudomonas fluorescens Pf0-1. P. Mosselii was first isolated and characterized in 2002, from clinical samples of hospitalized patient, but its association with the pathology remains unclear. We found that P. Mosselii ATCC BAA-99 and P. Mosselii MFY161 are cytotoxic towards Caco-2/TC7 cells, have low invasive capacity, and are able to alter the epithelial permeability of differentiated cells and damage the F-actin cytoskeleton. Their behavior resembles that of cytotoxic strains of P. Fluorescens. Moreover, their antibiotic resistance and potential role as shuttles for acquired beta-lactamases resistance suggest that P. Mosselii strains may be more studied. The cytotoxicity of P. Fluorescens Pf 0-1 was also tested and we found it very low in our experimental conditions. However, we were interested to notice that this bacteria possesses a translocator protein (TSPO) which is largely conserved during evolution (Eukarya, Archae and Bacteria), but which role was unknown for this bacteria. Then, our next work consisted in studying the expression of this protein and its potential function(s). We found that tspo is cotranscribed with an hybrid histidine kinase gene (Pfl01_2810) which is situated upstream to the tspo gene. This operon is transiently expressed during the growth of the bacteria in LB medium at 28°C. The transcriptional activity is modified with temperature, null at 18°C and highly increased at 32°C, suggesting a regulation by the sigma factor RpoH, generally activated in response to thermal stress. Hyperosmolarity (NaCl, Sucrose), or D-cycloserin enhanceds the transcriptional activity of the operon, and provoked a parietal stress and regulation of the sigma factor AlgU, via RpoH. Mitochondrial TSPO ligands as cholesterol, involved in membrane fluidity, also favors the expression of tspo, as well as other Eukaryotic ligands as PK11195 or protoporphyrin IX, suggesting a functional conservation between bacterial TSPO from P. Fluorescens Pf0-1 and its Eukaryotic ortholog
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Shen, Weiping. "Regulation of Escherichia coli pyrBI Gene Expression in Pseudomonas fluorescens." Thesis, University of North Texas, 1995. https://digital.library.unt.edu/ark:/67531/metadc278188/.
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Pseudomonas fluorescens does not appear to regulate the enzymes of de novo
pyrimidine biosynthesis at the level of gene expression. Little or no apparent repression of pyr gene expression is observed upon addition of exogenous pyrimidines to the growth medium. The Escherichia coli pyrBI genes for aspartate transcarbamoylase (ATCase) were sized down and cloned into the broad host range plasmid, pKT230. Upon introduction into a P.fluorescenspyrB mutant strain, ATCase showed repression in response to exogenously fed pyrimidine compounds. Thus, it was possible to bring about changes in pyrimidine nucleotide pool levels and in transcriptional regulation of gene expression at the same time.
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Teselkin, Oleksiy. "Repeatability of the Adaptation of Pseudomonas fluorescens to Low Glucose." Thèse, Université d'Ottawa / University of Ottawa, 2014. http://hdl.handle.net/10393/30986.
Full textAbstract:
Inspired by Gould, who claimed life would be arriving at a different outcome each time it were allowed to run from the same beginning, I have attempted to determine the repeatability of the adaptive course of one Pseudomonas fluorescens lineage. In addition, my study aimed to establish whether the likelihood of parallel evolution of the two synonymous single-nucleotide substitutions was contingent upon a prior motility-impairing deletion or a prior increase in fitness. Further, the study was designed to provide empirical data addressing the long-standing question of the effect of starting fitness on the ensuing rate of adaptation.
Although no exact replay of the initial evolutionary trajectory was observed, I have demonstrated that gtsB, but not gtsC gene, is likely to be a mutational hotspot under the low glucose with a recovery of two undescribed mutations in gtsB. My data are consistent with a notion that substitutions in gtsB may be contingent upon Δ35kB(fliJ-PFLU4466) motility-impairing deletion, but not the fitness increase associated with it.
Finally, the features of the adaptive landscape of P. fluorescens in the minimal glucose provide languid support for Fisher’s hypothesis of a decrease in adaptation rate with the rise in the starting fitness.
Taken together, these original results reinforce the non-negligible role of history in shaping the outcomes of biological evolution and call for caution in attempting a formulation of rigid predictive models of evolutionary change.
Inspiré par les travaux de Stephen J. Gould qui affirmait que la vie sur terre arriverait à une forme différente si elle repartait à zéro, je présente ici mes travaux où je teste la reproductibilité du cours adaptatif d’une lignée expérimentale de Pseudomonas fluorescens. L’objectif de cette étude était de déterminer si la probabilité que deux mutations synonymes évoluent en parallèle est affectée par la présence d’une délétion affectant la motilité de la bactérie ou de l’augmentation de la valeur sélective de celle-ci. De plus, le design expérimental de cette étude permet de tester si la valeur sélective initiale d’une population affecte le taux d’adaptation de cette même population.
Bien d’une reproductibilité exacte du cours adaptatif initial ne fut pas observée, je démontre que le gène gtsB est probablement un « hotspot »mutationnel permettant l’adaptation à de bas niveau de glucose, ayant trouvé deux mutations dans ce site; alors que le gène gtsC ne l’est pas. Mes données sont également conséquentes avec le fait que les mutation dans le gène gtsB dépendent de l’effet de la délétion Δ35kB(fliJ-PFLU4466) affectant la motilité de la bactérie, mais non de l’augmentation de la valeur sélective qui y est associée. Finalement, la forme du plateau adaptative associé à de bas niveaux de glucose chez P. fluorescens supporte l’hypothèse émise par Fisher qui stipule que le taux d’adaptation d’un organisme diminue avec la valeur sélective initiale qui y est associée.
L’ensemble de ces résultats supporte le rôle non-négligeable de l’histoire de vie d’une population en ce qui attrait à l’évolution future de cette même population. Aussi, ces résultats appelle à la prudence quand vient le temps de formuler des modèles prédictifs des changements évolutifs d’une population.
30
Dorr, Patrick Karl. "Cyanide oxygenase and cyanase activities of Pseudomonas fluorescens NCIMB 11764." Thesis, University of Kent, 1990. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.292714.
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Braithwaite, Kerynne Lindsay. "Novel plant cell wall hydrolases from Pseudomonas fluorescens subspecies cellulosa." Thesis, University of Newcastle Upon Tyne, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.294928.
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Gurney, Rachel. "Biosynthesis of the antibiotic mupirocin by Pseudomonas fluorescens NCIMB 10586." Thesis, University of Birmingham, 2013. http://etheses.bham.ac.uk//id/eprint/4204/.
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The mupirocin biosynthetic pathway belongs to the trans-AT group in which acyltransferase (AT) activity is provided by a separate polypeptide (MmpC) rather than in cis as found in the typical type I polyketide synthases. AT docking domains have been documented in trans-AT PKS clusters for ten years yet little functional evidence is available. The cluster shows many interesting features that must be understood to create novel products. Specificity studies demonstrated that AT2 performs the typical AT function of loading malonyl-CoA to ACPs throughout the cluster. Mutagenesis studies demonstrated the importance of AT active site residues for protein structural integrity, acquisition and transfer of malonate and propose an alternate role for AT1 as a proofreading enzyme responsible for hydrolysing truncated intermediates from the pathway. Consequently an edit, reload, reduce model for MmpC is proposed. Mutagenesis of docking domains led to a halt in mupirocin production and suggested that docking domains are required for structural integrity of the Mmps or for guiding the ACPs into the correct position for interactions with their respective partners. Studies involving a mutated ACP3 protein confirmed the importance of Trp55, as demonstrated by structural changes and the inability of the protein to accept malonate from AT2.
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Amin-Hanjani, Soheila. "Luminescence based detection of genetically modified Pseudomonas fluorescens in soil." Thesis, University of Aberdeen, 1992. http://digitool.abdn.ac.uk/R?func=search-advanced-go&find_code1=WSN&request1=AAIU043327.
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Methods currently available for the detection and enumeration of genetically modified micro-organisms in the environment include culturing methods, direct microscopic detection and nucleic hybridization techniques. The aim of this project was to develop luminescence as a molecular based-marker system in Pseudomonas fluorescens. The lux genes, originally isolated from Vibrio fischeri, were introduced into Ps. fluorescens on plasmid vectors and on the chromosome. The efficiency of these two strategies for the detection of Ps. fluorescens in soil was assessed. Luminometry was used to estimate biomass concentration during growth. The sensitivity of luminescence detection was greater for the plasmid marked Ps. fluorescens in both liquid culture and soil, however, cellular light output was less closely linked to biomass concentration. Enumeration of cells by luminometry was only possible for growing cells as light output is correlated with microbial activity. The lux chromosomal marker was stable in liquid culture for at least 200 generations and in soil for up to 135 days. The plasmid borne lux genes had a half-life of 20 generations in liquid culture. After inoculation in sterile soil, plasmid loss was only observed during cellular growth. The frequency of transfer of the lux genes from Ps. fluorescens, by conjugation and transformation, was assessed in liquid culture. Mobilisation of these genes by three self transmissible plasmids was negligible due to the instability of these vectors in this host. Transformation of Ps. stutzeri with lux genes, by cell contact, was at frequencies below levels of detection. Luminescence provided a valuable marker for tracking pseudomonads in soil. Detection of marked strains by luminometry provided a sensitive, rapid and non-extractive technique for enumeration of growing cells and measurement of microbial activity. As the chromosomally encoded lux genes were stable, regardless of growth conditions, and emitted sufficient levels of light to enable visual enumeration of colonies by eye, this was considered the best system for long term risk assessment studies.
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Chou, Chia-Ni. "Purification of Cyanide-Degrading Nitrilase from Pseudomonas Fluorescens NCIMB 11764." Thesis, University of North Texas, 2010. https://digital.library.unt.edu/ark:/67531/metadc33224/.
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Cyanide is a well known toxicant that arises in the environment from both biological and industrial sources. Bacteria have evolved novel coping mechanisms for cyanide and function as principal agents in the biosphere for cyanide recycling. Some bacteria exhibit the unusual ability of growing on cyanide as the sole nitrogen source. One such organism is Pseudomonas fluorescens NCIMB 11764 (Pf11764) which employs a novel oxidative mechanism for detoxifying and assimilating cyanide. A unique complex of enzymes referred to as cyanide oxygenase (CNO) is responsible for this ability converting cyanide to ammonia which is then assimilated. Because one component of the four member CNO complex was previously shown to act on cyanide independent of the other members, its characterization was sought as a means of gaining a better understanding of the overall catalytic mechanism of the complex. Preliminary studies suggested that the enzyme belonged to a subset of nitrilase enzymes known as cyanide dihydratases (CynD), however, a cynD-like gene in Pf11764 could not be detected by PCR. Instead, a separate nitrilase (Nit) linked to cyanide metabolism was detected. The corresponding nit gene was shown to be one of a conserved set of nit genes traced to a unique cluster in bacteria known as Nit1C. To determine whether the previously described CynD enzyme was instead Nit, efforts were undertaken to isolate the enzyme. This was pursued by cloning and expressing the recombinant enzyme and by attempting to isolate the native enzyme. This thesis is concerned with the latter activity and describes the purification of a Nit-like cyanide-degrading nitrilase (NitCC) from Pf11764 to ~95% homogeneity. Purification was greatly facilitated by the discovery that fumaronitrile, as opposed to cyanide, was the preferred substrate for the enzyme (20 versus 1 U/mg protein, respectively). While cyanide was less effective as a substrate, the specificity for cyanide far outweighed that (10,000 fold) of the recombinant enzyme (NitPG) implying that the native NitCC protein purified in this work is different from that of the cloned recombinant. Further evidence of this was provided by molecular studies indicating that the two proteins differ in mass (34.5 and 38 kDa, respectively) and amino acid sequence. In summary, two different Nit enzymes are encoded by Pf11764. While the two share greater than 50% amino acid sequence identity, the results suggest that the native NitCC enzyme purified in this work functions better as a cyanide-degrading nitrilase and is one of four enzyme components comprising CNO required for Pf11764 cyanide assimilation.
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Dal, Bello Elena <1983>. "Vanillin production from ferulic acid with Pseudomonas fluorescens BF13-1p4." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2013. http://amsdottorato.unibo.it/5482/1/PhD_Thesis_EDB.pdf.
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Bioconversion of ferulic acid to vanillin represents an attractive opportunity for replacing synthetic vanillin with a bio-based product, that can be label “natural”, according to current food regulations. Ferulic acid is an abundant phenolic compound in cereals processing by-products, such as wheat bran, where it is linked to the cell wall constituents. In this work, the possibility of producing vanillin from ferulic acid released enzymatically from wheat bran was investigated by using resting cells of Pseudomonas fluorescens strain BF13-1p4 carrying an insertional inactivation of vdh gene and ech and fcs BF13 genes on a low copy number plasmid. Process parameters were optimized both for the biomass production phase and the bioconversion phase using food-grade ferulic acid as substrate and the approach of changing one variable while fixing the others at a certain level followed by the response surface methodology (RSM). Under optimized conditions, vanillin up to 8.46 mM (1.4 g/L) was achieved, whereas highest productivity was 0.53 mmoles vanillin L-1 h-1). Cocktails of a number of commercial enzyme (amylases, xylanases, proteases, feruloyl esterases) combined with bran pre-treatment with steam explosion and instant controlled pressure drop technology were then tested for the release of ferulic acid from wheat bran. The highest ferulic acid release was limited to 15-20 % of the ferulic acid occurring in bran, depending on the treatment conditions. Ferulic acid 1 mM in enzymatic hydrolyzates could be bioconverted into vanillin with molar yield (55.1%) and selectivity (68%) comparable to those obtained with food-grade ferulic acid after purification from reducing sugars with a non polar adsorption resin. Further improvement of ferulic acid recovery from wheat bran is however required to make more attractive the production of natural vanillin from this by-product.
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Dal, Bello Elena <1983>. "Vanillin production from ferulic acid with Pseudomonas fluorescens BF13-1p4." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2013. http://amsdottorato.unibo.it/5482/.
Full textAbstract:
Bioconversion of ferulic acid to vanillin represents an attractive opportunity for replacing synthetic vanillin with a bio-based product, that can be label “natural”, according to current food regulations. Ferulic acid is an abundant phenolic compound in cereals processing by-products, such as wheat bran, where it is linked to the cell wall constituents. In this work, the possibility of producing vanillin from ferulic acid released enzymatically from wheat bran was investigated by using resting cells of Pseudomonas fluorescens strain BF13-1p4 carrying an insertional inactivation of vdh gene and ech and fcs BF13 genes on a low copy number plasmid. Process parameters were optimized both for the biomass production phase and the bioconversion phase using food-grade ferulic acid as substrate and the approach of changing one variable while fixing the others at a certain level followed by the response surface methodology (RSM). Under optimized conditions, vanillin up to 8.46 mM (1.4 g/L) was achieved, whereas highest productivity was 0.53 mmoles vanillin L-1 h-1). Cocktails of a number of commercial enzyme (amylases, xylanases, proteases, feruloyl esterases) combined with bran pre-treatment with steam explosion and instant controlled pressure drop technology were then tested for the release of ferulic acid from wheat bran. The highest ferulic acid release was limited to 15-20 % of the ferulic acid occurring in bran, depending on the treatment conditions. Ferulic acid 1 mM in enzymatic hydrolyzates could be bioconverted into vanillin with molar yield (55.1%) and selectivity (68%) comparable to those obtained with food-grade ferulic acid after purification from reducing sugars with a non polar adsorption resin. Further improvement of ferulic acid recovery from wheat bran is however required to make more attractive the production of natural vanillin from this by-product.
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Fields, Christopher J. "Comparative biochemistry and genetic analysis of nucleoside hydrolase in Escherichia coli, Pseudomonas aeruginosa, and Pseudomonas fluorescens." Thesis, University of North Texas, 2002. https://digital.library.unt.edu/ark:/67531/metadc3290/.
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The pyrimidine salvage enzyme, nucleoside hydrolase, is catalyzes the irreversible hydrolysis of nucleosides into the free nucleic acid base and D-ribose. Nucleoside hydrolases have varying degrees of specificity towards purine and pyrimidine nucleosides. In E. coli, three genes were found that encode homologues of several known nucleoside hydrolases in protozoa. All three genes (designated yaaF, yeiK, and ybeK) were amplified by PCR and cloned. Two of the gene products (yeiK and ybeK) encode pyrimidine-specific nucleoside hydrolases, while the third (yaaF) encodes a nonspecific nucleoside hydrolase. All three were expressed at low levels and had different modes of regulation.
As a comparative analysis, the homologous genes of Pseudomonas aeruginosa and P. fluorescens (designated nuh) were cloned. Both were determined to encode nonspecific nucleoside hydrolases. The nucleoside hydrolases of the pseudomonads exhibited markedly different modes of regulation. Both have unique promoter structures and genetic organization. Furthermore, both pseudomonad nucleoside hydrolase were found to contain an N-terminal extension of 30-35 amino acids that is shown to act as a periplasmic-signaling sequence. These are the first two nucleoside hydrolases, to date,that have been conclusively demonstrated to be exported to the periplasmic space. The
physiological relevance of this is explained.
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Selsaas, Eirik. "Karakterisering av gener som påvirker biosyntesen av alginat i Pseudomonas fluorescens." Thesis, Norges teknisk-naturvitenskapelige universitet, Institutt for bioteknologi, 2011. http://urn.kb.se/resolve?urn=urn:nbn:no:ntnu:diva-12753.
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Pseudomonas fluorescens har evne til å produsere store store mengder alginat. Tidligere er det laget en stamme, P. fluorescens SBW25 MS1 ΔalgC::TnKb61, hvor algD-operonet og algC er satt under kontroll av Pm-promotor. Dette gjør at den produserer alginat ved tilsats av m-toluensyre. Denne stammmen har blitt mutert med transposon og effekten på alginatproduksjonen har blitt undersøkt. Ved transposonmutagenese ble det funnet 180 mutanter med endret alginatproduksjon, og insersjonspunktet for transposonet ble identifisert.I to av disse mutantene var henholdsvis genene clpA og PFLU3927 inaktivert. clpA er en Clp-protease, mens PFLU3927 er en protease som tilhører lon-proteasefamilien. I denne oppgaven skulle disse genenes rolle i biosyntesen av alginat verifiseres. For å gjøre dette ble mutantene komplementert ved å sette inn villtypevarianten av genet inn i mutanten. Innføringen av genet ble gjort ved bruk av transposon. Siden algD-operonet var satt under kontroll av Pm-promotoren i stammen som ble benyttet, skal denne stammen være uavhengig av sigmafaktoren AlgU, som normalt er nødvendig for ekspresjon av algD-operonet. For å teste om stammen virkelig var uavhengig av AlgU, ble det laget en mutant hvor AlgU ble inaktivert. Dette ble gjort ved å fjerne en del i algU ved bruk av PCR. Videre ble villtypegenet erstattet med delesjonsgenet i P. fluorescens. Dette ble gjort ved bruk av homolog rekombinering.For å se hvilken effekt genene hadde på alginatproduksjonen, ble det målt alginatproduksjon i transposonmutantene og i de komplementerte stammene som ble laget i dette arbeidet. Alginatmålingene gikk ikke som ventet, da det ikke kunne registreres alginat i de positive kontrollene. Det var kun to stammer hvor det ble registrert alginatproduksjon, 30O1 clpA og 30O1 clpA::TnES4. Dette forsøket ble utført gjentatte ganger, men med samme resultat. Det er foreslått at forskjellen i alginatproduksjon mellom mediene PIM og DEF3 i tidligere forsøk på clpA kan komme av forskjeller i fosfatmengde og/eller peptonmengde i mediene. Det er også foreslått at PFLU3927 kan være et varmesjokkprotein og at proteinaktiviteten kan økes av ulike former for stress, og videre at dette kan føre til at alginatproduksjonen endres som følge av det.
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Burlinson, Peter. "Characterization and regulation of anti-eukaryote factors in Pseudomonas fluorescens. NZI7." Thesis, University of Oxford, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.491327.
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This study examined Pseudomonas 'NZ strains' that cause blotch disease on the commercial mushroom Agaricus bisporus. Mushroom-pathogenic Pseudomonas represent an interesting example of bacteria associated with a fungal host system. The mycosphere also contains other organisms that Pseudomonas may interact with, thus strains isolated from this environment may also contribute towards our understanding of the molecular basis of opportunistic pathogenesis and bacterial survival in nature. Phylogenetics was used to assess the relatedness of NZ strains and their position in the Pseudomonas genus. It was hypothesised that NZ strains may possess means to resist predators and parasitize other eukaryotes present in the mycosphere. Evidence for this was sought by assessing their interactions with several fungi, the nematode Caenorhabditis elegans and the social amoeba Dictyostelium discoideum. The differential interactions observed suggest some strains have specific mechanisms to antagonize bacteriovores. NZ strains were surveyed for production of potentially antagonistic exofactors and also for possession of type III secretion systems, which were identified in several strains. The phylogenetic distribution of these factors suggests traits such as cyanide production, exochitinase activity and type III secretion may be restricted to certain P.f1uorescens lineages. Transposon mutagenesis of strain NZI7 showed production of the toxin tolaasin was essential for production of brown blotch symptoms but appeared not to influence C.elegans or D.discoideum interactions. Cyanide production influenced both A.bisporus and C.elegans interactions. A novel screening procedure identified NZI7 mutants with decreased resistance to C.elegans feeding and identified a gene cluster hypothesized to produce a metabolite contributing to the NZI7 inedibility phenotype. The expression of these genes was characterised using luxCDABE transposon reporter mutants and the global sensor kinase GacS was found to control their expression. GacS was also found to regulate the production of anti-fungal factors in strain NZ011.
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Melnyk, Anita H. "Adaptive landscapes in evolving populations of Pseudomonas fluorescens in simple environments." Thesis, University of Ottawa (Canada), 2010. http://hdl.handle.net/10393/28876.
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The adaptive landscape heuristic can be used to answer the question "how predictable is evolution?" because its topology will impact the repeatability of evolution. In my Masters research I addressed this question in two ways: (1) I reviewed empirical adaptive landscape studies in the fields of directed protein evolution and microbial experimental evolution and (2) I performed a selection experiment to characterize adaptive landscape topology by measuring variance in fitness and metabolic phenotype within and among genetically distinct Pseudomonas fluorescens strains in two environments. Empirical studies have found that protein level landscapes are generally smooth, however, population level landscapes are rugged even in simple environments. Experimentally I found that the pattern of variance in fitness and metabolic phenotype was unique to the selection environment. The response to selection was highly repeatable at the level of fitness, but the underlying genetic routes taken were different for each environment and more variable in xylose than in glucose, suggesting a more rugged underlying landscape. More generally, my research suggests that making statements about the predictability of adaptive evolution at the population level may be challenging and wi11likely depend on the specifics of the environment in which selection occurs.
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McCarthy, Conor Neil, and n/a. "Regulatory Elements Controlling Lipase and Metalloprotease Production in Pseudomonas fluorescens B52." Griffith University. School of Health Science, 2003. http://www4.gu.edu.au:8080/adt-root/public/adt-QGU20031015.124744.
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Psychrotrophic bacteria, such as Pseudomonas fluorescens B52, are a major cause of milk spoilage at refrigeration temperature due to the production of lipolytic and proteolytic enzymes. Regulatory mechanisms controlling the production of lipase and protease by the B52 lipA and aprX genes were investigated. Transposon mutagenesis identified the possible involvement of a poly-A polymerase enzyme which destabilises mRNA by 3' polyadenylation. A homologue of the E. coli EnvZ/OmpR two-component sensor/regulator system was identified by transposon mutagenesis and shown to repress lipase and protease production. This system responds to Na+ and K+ concentration in E. coli and these ions were also shown to repress lipase and protease expression in B52, however the EnvZ/OmpR system is not solely responsible for this. Assays of translational lacZ fusions with aprX and lipA were used to speculate on the mechanism by which Na+ and EnvZ/OmpR repress the aprX-lipA operon. A membrane-bound sensor, MspA, which regulates protease production in P. fluorescens LS107d2, was shown to exist in B52 but mutagenesis of the B52 mspA gene had no effect on lipase and protease expression. A homologue of the P. fluorescens CHA0 rsmA gene, encoding an RNA-binding translation repressor, was found in B52. Although aprX and possibly lipA contain consensus sequences for RsmA, mutagenesis of rsmA had no significant effect on lipase and protease expression. Repression of lipase and protease expression by Na+ was increased by expression of the P. fluorescens M114 pbrA sigma-factor gene in B52.
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Shields, Jennifer Ann. "Biosynthesis of the polyketide antibiotic mupirocin by Pseudomonas fluorescens NCIMB 10586." Thesis, University of Birmingham, 2008. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.506425.
Full textAbstract:
A 75 kb gene cluster in Pseudomonasfl uorescens NCIMB 10586 encodesa modular polyketide synthase responsible for biosynthesis of the antibiotic mupirocin. All seven PKS modules lack intrinsic acyl transferase (AT) domains. Two AT domains within the multifunctional polypeptide MmpC are thought to operate in trans. An unusually large tailoring region encodes 26 discrete proteins that modify the polyketide/fatty acid backbone, in addition to providing resistance and regulatory functions. This study investigated the functions of four tailoring region genes. MupJ, MupK and mAcpC were revealed to be involved in insertion of a β-methyl group, while MupN was shown to have phosphopantetheinyl transferase activity. Five tailoring acyl carrier proteins (mAcps) were found to be specific and non-interchangeable, and the influence of C-termini on mAcp-specificity was investigated. Attempts to overexpress mAcps in P. fluorescens revealed that a broad-host-range IncP-1 vector is unstablein this host,p romptingt he constructiono f a new IncP-9v ector for use in P. fluorescens. The importance of the two ATs and the third domain of MmpC were demonstrated by mutagenesis and complementation. Overexpression of AT domains did not increase mupirocin production but specific heterologous trans-AT domains could malonylate the mupirocin PKS.
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Ferreira, Luis Manuel dos Anjos. "Molecular analysis of cellulases and xylanses from Pseudomonas fluorescens subspecies cellulosa." Thesis, University of Newcastle Upon Tyne, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.316083.
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Kellett, Louise Elizabeth. "The molecular biology of xylanolytic enzymes from Pseudomonas fluorescens subsp. cellulosa." Thesis, University of Newcastle Upon Tyne, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.315636.
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Zahran, Ahmed Shawky. "Production and properties of a protease secreted by Pseudomonas fluorescens R8." Thesis, University of Edinburgh, 1988. http://hdl.handle.net/1842/11965.
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Wilkins, Annekathrin. "Factors influencing the dispersal of Pseudomonas fluorescens NZI7 by Caenorhabditis elegans." Thesis, University of Oxford, 2016. https://ora.ox.ac.uk/objects/uuid:6bf58183-f197-490d-86d4-633ae8d46c06.
Full textAbstract:
Caenorhabditis elegans is a natural predator of the mushroom pathogen Pseudomonas fluorescens NZI7. The bacterial mechanisms for reducing predation by the nematode through the secretion of secondary metabolites have been described, but not yet fully explored. The behaviour of nematodes is influenced by the different factors produced by the pseudomonads. In this thesis we develop a range of assays to link the behaviour of C. elegans to these factors to identify their role in bacteria-nematode interactions. We show that these factors play two distinct roles: they may either repel nematodes, or harm them. This permits the classification of mutants of P. fl. NZI7 lacking these factors as either attractive, edible or both. Many studies of C. elegans behaviour have demonstrated that the nematode can distinguish between different food sources. Our results show two distinct types of response: chemotaxis drives the response to attractive or repellent stimuli, and nematodes also show a choice behaviour that is independent of chemotaxis. This choice behaviour is determined by bacterial edibility and requires nematodes to come into contact with the bacteria. This contact is the foundation of the bacterial dispersal by nematodes. By making use of the luminescence property of the available bacterial mutants, we demonstrate an intimate link between the behaviour of C. elegans and the success with which bacteria are disseminated: if nematodes are induced to regularly leave a bacterial colony, whether through their genotype or the low edibility of the food, then they will spread bacteria effectively. Throughout this thesis, we use computational simulations based on a hybrid cellular automaton model to represent the nematode-bacteria interactions. These simulations recreate the observed behaviour of the system, thus they help to confirm our hypotheses and establish the fundamental aspects of the interactions between the two species.
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Parab, Preeti. "Requirements for Cell-Free Cyanide Oxidation by Pseudomonas Fluorescens NCIMB 11764." Thesis, University of North Texas, 2000. https://digital.library.unt.edu/ark:/67531/metadc2614/.
Full textAbstract:
The involvement of cyanide oxygenase in the metabolism of pyruvate and a-ketoglutarate-cyanohydrin was investigated and shown to occur indirectly by the consumption of free cyanide arising from the cyanohydrins via chemical dissociation. Thus, free cyanide remains the substrate, for which the enzyme displays a remarkably high affinity (Kmapp,4 mM). A model for cyanide utilization is therefore envisioned in which the substrate is initially detoxified by complexation to an appropriate ligand followed by enzymatic oxidation of cyanide arising at sublethal levels via chemical dissociation. Putative cyanide oxygenase in cell extracts consumed both oxygen and NADH in equimolar proportions during cyanide conversion to CO2 and NH3 and existed separately from an unknown heat-stable species responsible for the nonenzymatic cyanide-catalyzed consumption of oxygen. Evidence of cyanide inhibition and nonlinear kinetics between enzyme activity and protein concentration point to a complex mechanism of enzymatic substrate conversion.
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McCarthy, Conor Neil. "Regulatory Elements Controlling Lipase and Metalloprotein Production in Pseudomonas fluorescens B52." Thesis, Griffith University, 2003. http://hdl.handle.net/10072/367432.
Full textAbstract:
Psychrotrophic bacteria, such as Pseudomonas fluorescens B52, are a major cause of milk spoilage at refrigeration temperature due to the production of lipolytic and proteolytic enzymes. Regulatory mechanisms controlling the production of lipase and protease by the B52 lipA and aprX genes were investigated. Transposon mutagenesis identified the possible involvement of a poly-A polymerase enzyme which destabilises mRNA by 3' polyadenylation. A homologue of the E. coli EnvZ/OmpR two-component sensor/regulator system was identified by transposon mutagenesis and shown to repress lipase and protease production. This system responds to Na+ and K+ concentration in E. coli and these ions were also shown to repress lipase and protease expression in B52, however the EnvZ/OmpR system is not solely responsible for this. Assays of translational lacZ fusions with aprX and lipA were used to speculate on the mechanism by which Na+ and EnvZ/OmpR repress the aprX-lipA operon. A membrane-bound sensor, MspA, which regulates protease production in P. fluorescens LS107d2, was shown to exist in B52 but mutagenesis of the B52 mspA gene had no effect on lipase and protease expression. A homologue of the P. fluorescens CHA0 rsmA gene, encoding an RNA-binding translation repressor, was found in B52. Although aprX and possibly lipA contain consensus sequences for RsmA, mutagenesis of rsmA had no significant effect on lipase and protease expression. Repression of lipase and protease expression by Na+ was increased by expression of the P. fluorescens M114 pbrA sigma-factor gene in B52.Thesis (PhD Doctorate)
Doctor of Philosophy (PhD)
School of Health Sciences
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Careli, Roberta Torres. "Adesão de Pseudomonas fluorescens em superfícies utilizadas no processamento de alimentos." Universidade Federal de Viçosa, 2005. http://www.locus.ufv.br/handle/123456789/9091.
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Conselho Nacional de Desenvolvimento Científico e Tecnológico
A adesão de Pseudomonas fluorescens ATCC 13525 foi avaliada pela microscopia de epifluorescência (EPF) e contagem padrão em placas (CPP) em superfícies usadas no processamento de alimentos nos tempos de contato 0, 2, 4, 6, 8 e 10 h. O aumento da adesão entre as superfícies, em razão do tempo, não foi acompanhado de forma similar pelas técnicas avaliadas. Por exemplo, no tempo quatro horas, as superfícies que apresentaram maiores logaritmos de UFC.cm-2 e que não apresentaram diferença significativa na adesão pelo teste de Scott-Knott (P > 0,05) foram poliuretano rugoso dupla face, silicone revestido com tecido, poli (cloreto de vinila) revestimento grosso com tecido, granito e mármore pela técnica EPF. Já pela técnica CPP, no mesmo tempo de contato, os maiores logaritmos de UFC.cm-2 foram para superfícies de silicone revestido com tecido, poliuretano rugoso dupla face, granito e poliuretano revestido com tecido. As superfícies de mármore, granito, poli (cloreto de vinila) revestimento grosso com tecido, poliuretano rugoso dupla face e silicone revestido com tecido diferiram das demais no grau de adesão, expresso em UFC.cm-2 (P < 0,05) nos tempos 4, 6, 8 e 10 horas, quando avaliadas pela técnica da epifluorescência, e de 2 e 10 horas, quando avaliadas pela contagem padrão em placas. Numa outra forma de avaliação, constatou-se também a diferença entre as 12 técnicas com relação ao agrupamento das superfícies de acordo com a similaridade de adesão. Assim, as superfícies que apresentaram maior percentagem de similaridade e maior média geral com relação à adesão pela técnica EPF foram mármore, granito e poliuretano rugoso dupla face. No caso da CPP, este mesmo fato foi constatado com as superfícies de poliuretano rugoso dupla face, silicone revestido com tecido e granito. As superfícies apresentaram características de microtopografias muito diferentes quando observadas por microscopia eletrônica de varredura, o que pode justificar as diferenças entre os graus de adesão observados. As técnicas possuem comportamentos diferentes para cada tempo de contato. Constatou-se que a CPP, além de fornecer resultados menores do que a EPF, também permite a detecção de valores de adesão mais baixos, sendo considerada uma técnica mais sensível. Porém, a CPP fornece resultados mais demorados do que a EPF. Para que haja detecção pela EPF, a quantidade de células aderidas aos cupons deve estar com contagens médias de uma célula por campo, no mínimo. A EPF permite verificar a morfologia das células, bem como, a distribuição destas bactérias aderidas às diferentes superfícies avaliadas. É recomendável utilizar a EPF para a quantificação de bactérias sésseis, principalmente quando as contagens sejam menores ou iguais a 4,1 x 105 UFC.cm-2. Este experimento mostrou que Pseudomonas fluorescens ATCC 13525 aderiu nas superfícies avaliadas. Entretanto, não há como sugerir a superfície mais recomendada para a utilização em processamento de alimentos devido a suas aplicações específicas. Os resultados mostram a importância de práticas higiênicas corretas na indústria de alimentos.
The adhesion of Pseudomonas fluorescens ATCC 13525 was evaluated by the epifluorescence microscopy (EPF) and the plate count method (CPP) to surfaces used in food processing at 0, 2, 4, 6, 8 and 10 h contact times. The adhesion increase among the surfaces, in relation to time, was not followed in a similar way by the evaluated techniques. For example, in four hours, the surfaces which showed greater CFU.cm-2 logarithms and that did not show a significant difference in the adhesion by the Scott- Knott test (P > 0.05) were the double-faced rugous polyurethane, silicon coated with cloth, PVC thick coated with cloth, granite and marble by the EPF technique. Whereas by the CPP technique, in the same contact time, the greater CFU.cm-2 logarithms were for the silicone coated with cloth, double-faced rugous polyurethane, granite and polyurethane coated with cloth surfaces. The marble, granite, PVC thick coated with cloth, double faced rugous polyurethane and silicone coated with cloth surfaces differed form the others in the adhesion degree expressed in CFU.cm-2 (P > 0.05) in the 4, 6, 8 and 10 h times, when evaluated by the epifluorescence technique and, at 2 and 10 h, when evaluated by the plate count method. In another kind of evaluation, the differences between the techniques concerning the surfaces cluster according to the adhesion similarity were also observed. Thus, the surfaces, which showed a greater similarity percentage and a greater general average concerning the adhesion by the 14 EPF technique, were marble, granite and double-faced rugous polyurethane. In the CPP, this same fact was observed with the double-faced rugous polyurethane, silicone coated with cloth and granite surfaces. The surfaces showed very different microtopography characteristics when observed by the scanning electron microscopy, which can justify the differences between the observed adhesion degrees. The techniques have different behaviors for each contact time. It was observed that CPP, besides providing results lower than the EPF, also allowed the detection of lower adhesion values, being considered a more sensitive technique. However, the CPP provides longer results than the EPF. So that there is a detection by EPF, the quantity of adhered cells to the coupons must be with, at least, an average count of one cell per field. The EPF allows the cell morphology assay as well as the distribution of these bacteria adhered to the different evaluated surfaces. It is recommendable to use the EPF technique for the sessile bacteria quantifying, especially when the count is lower or equal to 4,1 x 105 CFU.cm-2. This study showed that the Pseudomonas fluorescens ATCC 13525 adhered to the evaluated surfaces. However, there is no way to suggest the most recommendable surface for the use in the food processing due to their specific application. The results show the importance of the correct hygienic practices in the food industry.
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Andreani, Nadia Andrea. "INTO THE BLUE: Spoilage phenotypes of Pseudomonas fluorescens in food matrices." Doctoral thesis, Università degli studi di Padova, 2016. http://hdl.handle.net/11577/3424342.
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Spoilage induced by Pseudomonas strains is commonly found in a wide range of food products as a result of the ubiquitous presence of these strains and their ability to induce alteration through different mechanisms. Particular attention has been recently paid on those P. fluorescens strains able to induce a blue discolouration on several food matrices (e.g. dairy or meat products). Actually, poor data are available about this curious event that draw the attention of European consumer from 2010.
In the present manuscript a step-by-step investigation of the spoilage potential of Pseudomonas fluorescens species complex strains is reported, focusing in particular on the ability to produce an unpleasant blue pigment in food.
Firstly, some general information is given to the reader to understand the P. fluorescens group as food spoiler. Then, the application of a polyphasic approach is described with the aim to investigate 136 Pseudomonas fluorescens group strains. Additionally, the achievement and the analyses of draft genomes and transcriptomes for 4 P. fluorescens strains are described to investigate the biosynthetic pathways involved in the blue pigment production. The attempt to chemically characterise the blue molecule using MALDI-TOF mass spectrometry is also reported. Finally, the execution of a transposon-mediated mutagenesis is described to confirm previously obtained genomic data and to highlight further genes involved in the blue-pigment production.
The phenotypic and genotypic characterisation, based on the combination of classical microbiological tests and a MLST scheme, allowed the reconstruction of phylogenetic relationships among the isolates and the identification of a monophyletic group (named “the blue branch”) grouping all the blue-pigmenting and few uncoloured strains. The real involvement of these strains in the blue mozzarella event was confirmed by their ability to induce a blue discolouration on mozzarella cheese during a challenge test.
The genomic investigation confirmed the strict phylogenetic relationship between the strains belonging to the “blue branch”. Additionally, comparative genomic tools revealed the presence of a genetic cluster unique to the blue pigmenting strains containing a second copy of five trp genes, clearly involved in the blue pigment production. The biochemical characterisation of the pigment, hampered by strong issues of solubility, led to the conclusion that the molecule is an indigo-derivative.
Transposon-induced mutants confirmed the involvement of the previously identified unique cluster and the association of several genes affecting directly or indirectly the blue molecule production. Furthermore, the phenotypic characterisation of the mutants revealed a key role of iron in the production of the pigment, such as absence of any advantage of the wild-type strain in co-culture with a non-pigmenting mutant.
To conclude, the present work represents an exhaustive investigation of the spoilage potential of the blue-pigmenting P. fluorescens strains, giving to food industry reliable approaches to identify, track and prevent spoilage related to the growth of these interesting bacteria.Le alterazioni causate da ceppi di Pseudomonas sono solitamente riscontrate in una grande varietà di alimenti a causa del loro essere ubiquitari e dalla loro capacità di indurre modificazioni organolettiche negli alimenti mediante diversi meccanismi. Particolare attenzione è stata posta su alcuni ceppi di P. fluorescens in grado di indurre una colorazione blu in diverse matrici alimentari (quali prodotti lattiero-caseari o carne). In realtà, poche informazioni sono ad oggi disponibili riguardo al curioso caso che ha attirato l’attenzione pubblica a partire dal 2010. In questo lavoro è riportata un’analisi a più livelli del potenziale alternate dei ceppi appartenenti allo Pseudomonas fluorescens species complex, ponendo particolare attenzione alla capacità di produrre un indesiderato pigmento blu negli alimenti. In primo luogo, ai lettori sono date delle informazioni generali per una migliore comprensione di P. fluorescens come alterante alimentare. In seguito, è descritta la messa a punto e applicazione di un approccio polifasico con l’obbiettivo di indagare 136 ceppi appartenenti al gruppo P. fluorescens. Inoltre, sono descritti l’ottenimento e le analisi dei genomi draft e dei trascrittomi di 4 ceppi di P. fluorescens con la finalità di comprendere il pathway biosintetico coinvolto nella produzione del pigmento blu. In aggiunta, è riportato il tentativo di caratterizzare chimicamente il pigmento mediante la metodica della spettrometria di massa MALDI-TOF. Infine, è riportata l’esecuzione della mutagenesi random con la finalità di confermare i risultati genomici precedentemente ottenuti e di individuare ulteriori geni coinvolti nella produzione del pigmento blu. La caratterizzazione fenotipica e genotipica, basata sulla combinazione di metodiche di microbiologia classica e di uno schema MLST, ha permesso la ricostruzione delle relazioni filogenetiche tra gli isolati e l’identificazione di un gruppo monofiletico (chiamato “ramo blu”) che raggruppa tutti i ceppi pigmentanti e pochi ceppi non-pigmentanti. Il reale coinvolgimento dei ceppi blu nei casi di mozzarella blu è stato confermato dalla possibilità degli stessi di indurre un’anomala colorazione blu su mozzarella durante un challenge test. Le analisi genomiche hanno confermato la stretta vicinanza filogenetica tra i ceppi del “ramo blu”. Inoltre, analisi di genomica comparativa hanno rivelato la presenza di un cluster genico unicamente presente nei ceppi blu, contenente una seconda copia di cinque dei sette geni per la biosintesi del triptofano, chiaramente coinvolto nella produzione del pigmento blu. La caratterizzazione biochimica del pigmento, resa difficoltosa da problemi di solubilità, ha portato alla conclusione che la molecola blu sia un derivato dell’indigo. I mutanti ottenuti mediante l’applicazione di trasposoni hanno confermato il coinvolgimento del cluster genico precedentemente identificato nella produzione del pigmento e l’associazione di ulteriori geni che influenzano direttamente o indirettamente la produzione della molecola blu. Inoltre, la caratterizzazione dei mutanti ha rivelato il ruolo importante del ferro nella produzione del pigmento e l’assenza di un effettivo vantaggio del ceppo wild-type posto in co-cultura con un mutante non pigmentante. In conclusione, questo studio rappresenta un’indagine esaustiva del potenziale alterante dei ceppi blu, dando inoltre all’industria alimentare sistemi efficaci per identificare, tracciare e prevenire l’alterazione indotta da questi interessanti ceppi.