Les Streptomyces sont des bactéries communes des écosystèmes forestiers tempérés. Elles sont présentes au sein de différentes niches écologiques telles que la rhizosphère des plantes, la mycorhizosphère ou encore la minéralosphère. Dans ces niches, elles interagissent avec de nombreux autres genres bactériens et différents champignons symbiotiques, pathogènes et saprophytes. Nous avons entrepris une étude sur l'interaction entre deux souches bactériennes modèles du sol Streptomyces ambofaciens ATCC23877 et Pseudomonas fluorescens BBc6R8. Des expériences de cocultures sur différents milieux gélosés ont révélé que chaque souche bactérienne affecte la production de métabolites secondaires chez l'autre partenaire. Ainsi, P. fluorescens inhibe la capacité de S. ambofaciens à produire la kinamycine, un « effet ping-pong » a été mis en évidence. D'autre part, sur un milieu déficient en fer, P. fluorescens est capable d'utiliser les desferrioxamines et la coelicheline produites par S. ambofaciens en tant que xénosiderophores et ne produit alors plus ses propres sidérophores, la pyoverdine et l’énantio-pyochéline. Enfin, au cours des études d’interactions, P. fluorescens BBc6R8 s’est révélée capable d’inhiber la production du pigment bleu diffusible γ-actinorhodine chez Streptomyces coelicolor A3(2) M145 en acidifiant le milieu par la production d’acide gluconique. L’ensemble de ces résultats nous permet de proposer différentes hypothèses sur l’importance écologique de ces trois interactions en les replaçant au sein du contexte de l’écosystème sol / Streptomyces are common bacteria in temperate forest ecosystems, which inhabit various ecological niches such as plant rhizosphere and mycorrhizosphere, as well as mineralosphere. In these niches, they interact with many other bacterial genera and different symbiotic, pathogenic and saprotrophic fungi. We have initiated a study on interaction between two soil model bacteria strains Streptomyces ambofaciens ATCC23877 and Pseudomonas fluorescens BBc6R8. Cocultures experiments on different agar media have revealed that each bacteria affects the production of secondary metabolites in the other partner. Thus, P. fluorescens inhibits the ability of S. ambofaciens to produce kanamycin, a "ping-pong effect" was highlighted. On the other hand, on iron deficient medium, P. fluorescens is able to use the desferrioxamines and coelichelin produced by S. ambofaciens as xenosiderophores and does no longer produce its own siderophores, pyoverdin and enantio-pyochelin. In another way, during pairwise co-culture experiments, Pseudomonas fluorescens BBc6R8 was shown to prevent the production of the diffusible blue pigment antibiotic γ-actinorhodin by Streptomyces coelicolor A3(2) M145 by acidifying the medium through the production of gluconic acid. Other fluorescent Pseudomonas strains and the opportunistic pathogen Pseudomonas aeruginosa PAO1 also prevented the γ-actinorhodin production in a similar way. Altogether these results prompt us to propose some hypotheses on the ecological significance of these three interactions by replacing them in the soil ecosystem context

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.

Mitt arbete handlar om mötet mellan rysk konstruktivism och science fiction. Detta har jag gestaltat i ett textilt rum inspirerad av science fictionkarraktären Doktorn ur Doctor Who. Doktorn är en utomjording av en långt mer utvecklad art och från en mer avancerad civilisation än människorna, som reser i tid och rum i sitt tids-och rymdskepp TARDIS. Jag har designat ett rum som är min textila tolkning av TARDIS innersta. Arbetet kretsar kring tankar om gestaltning av en flerdimensionalitet som bara kan anas, inte ses eller förstås. Jag har arbetat med optiska och optiskt omöjliga mönster, UV-ljus och fluorescens för att få en känsla av fler dimensioner än våra vanliga mänskliga fyra. Syftet är att förvirra betraktaren, att irritera ögat och hjärnan till att leta efter lösningar som inte finns. My thesis is about the meeting of russian constructivism and science fiction. I have constructed a textile room inspired by a fictional character, The Doctor from Doctor Who. The Doctor is an alien from an ancient and advanced civilization, travelling through time and space in his space-time ship TARDIS. The thesis is my textile interpretation of TARDIS’ center. I have studied the possibilities of creating dimensions, or the experience of dimensions, that humans can’t see or understand. For this I have studied optical patterns in combination with black lights and fluorescence. My aim is to confuse and irritate the eye and the brain of the beholder, and to make her look for solutions that aren’t there. / <p>Program: Textildesignutbildningen</p>

Fluorescens In Situ Hybridisering (FISH) är en cytogenetisk teknik som kan detektera genetiska sjukdomar och avvikelser i Deoxyribonukleinsyra (DNA) och Ribonukleinsyra (RNA). FISH börjar med kromosomutvinning av önskat analyspreparat, därefter får en direkt- eller indirekt fluorescensinmärkt probe (15-30 baspar lång) binda in till sin genetiska målsekvens via hybridisering. Preparatet kan sedan analyseras i fluorescensmikroskop för att bedöma om proben bundit in till sin målsekvens. I kursen ”Fördjupad laboratoriemetodik” för Biomedicinska analytikerprogrammet, Linnéuniversitetet utförs en FISH-laboration där resultaten varit otydliga och ej reproducerbara. Syftet med examensarbetet var att förbättra kurslaboration FISH som ges under kursen ”Fördjupad laboratoriemetodik”. Adherenta celler odlades till ≥ 3 x 106 i antal och till viss konfluens; primära endotelcell-linjen HCMEC till konfluenserna 60 % och 80 % och cancercell-linjerna VMM1 och H1915 till 80 % konfluens. Därefter skördades cellerna och dess respektive kromosomer utvanns. Kromosomerna undersöktes sedan med metoderna G-bandsfärgning, DNA-FISH och Multicolor-FISH (24X-probe) för tydliga och reproducerbara resultat. G-bandsfärgningen av HCMEC visade många hela interfasceller och få fria kromosomer för celler i 60 % konfluens. Både G-bandsfärgningen och DNA-FISH visade att HCMEC odlade till 80 % konfluens visade fria kromosomer från celler i metafas (celldelningsfas) där det fanns en svag signal för X-kromosomen. Multicolor-FISH-analys av VMM1 och H1915 gav tydliga resultat i fluorescensmikroskop där fria kromosomer var Multicolor-probeinmärkta; blå/aqua, röd och grön. Konklusionen är att vid kromosomutvinning från odlade adherenta celler bör dessa vara 80 % konfluenta. Detta för att ge tydliga och reproducerbara probeinfärgningar av kromosomer i metafas vid analys med Multicolor-FISH. Analys av 80 % konfluenta celler och användning av Multicolor-FISH-analys är en klar förbättring av kurslaborationen. / Fluorescens in situ hybridization (FISH) is used to detect cytogenetic aberrations and abnormalities of Deoxyribonucleic acid (DNA) and Ribonucleic Acid (RNA). The FISH begins with chromosome extraction of the desired cell preparation then a direct or indirect fluorescently labeled probe (15-30 base pair long) is hybridized to its genetic target sequence. The preparation can thereafter be analyzed in fluorescence microscope to see bound probe at chromosome level. In the course “Advanced laboratory methodology” for the Biomedical Scientist program, Linnaeus University, a FISH laboratory experiment is conducted where results have not been clear nor reproducible. The aim of this study was to improve the laboratory experiment FISH. Human Cardiac Microvascular Endothelial Cells (HCMEC) was grown to 60 % and 80 % confluence, to an estimated number of ≥ 3 x106, and analyzed by G-band staining and DNA-FISH. G-band staining showed many cells in interphase and few free chromosomes of cells with 60 % confluence. G-band staining and DNA-FISH showed that cells grown to 80 % confluence showed more free chromosomes from metaphase. The cancer cell lines VMM1 and H1915 were therefore grown to 80 % confluence and ≥ 3 x106. Multicolor-FISH on VMM1 and H1915 showed results from all painting probes blue/aqua, red and green. The conclusion is that in chromosomal extraction from cultured adherent cells should be 80 % confluent to give clear and reproducible probe staining of chromosomes in metaphase when assayed with Multicolor FISH. Analysis of 80 % confluent cells and the use of Multicolor FISH technology is a clear improvement to the “Advanced laboratory methodology” course.

The loss of rice plants (Oryza sativa L.) due to damage caused by abiotic stresses (water deficit and allelopathy) are recurrent, the objective was to evaluate the behavior of rice plants through agronomic and morphological changes under different layers of water in the soil (LAS) of 100%, 70%, 50% and 30% of field capacity (CC), and to identify the anatomical and physiological changes in plant in LAS 100% and 50% of CC, Induced by PGPR (Burkholderia pyrrocinia BRM-32113 and Pseudomonas fluorescens BRM-32111), that indicate the mitigating effect of the damages caused by the water deficit. And in plantings with residue of rice plants in the soil was to identify and understand the effect of the application of rhizobacteria on highland rice plants in consecutive plantations. Test one, The PGPR were subjected to temperature abiotic stresses (30, 35, 40 oC), salinity (0.5%, 7.5%), and water deficit (0, -0.2, - 0.4, -0.6, -0.8, -1.0, -1.2 Mpa) simulated by PEG with 10 replicates and evaluated the growth. Test two, the rice plants inoculated with: BRM-32111, BRM-32113 and control, were submitted the LAS of: 100, 70, 50 and 30% of CC, with three pots per treatment and five plants per pot. were evaluated: evapotranspirated water and water potential (ψam), biomass and length of plant and root, leaf area and relative chlorophyll content. Test three, plants Control, inoculated with BRM-32111 and BRM-32113, submitted to 100% and 50% of CC, and maintained for 28 days, and evaluated growth, physiology and anatomy. Test four, were used four treatments consisting of seed rice inoculated with P. fluorescens BRM-32111 in soil with residue, B. pyrrocinia BRM-32113 in soil with residue, plants control in soil with residue (CR)and plants control on soil without residue (SR) roots of plants rice (residues of Allelochemicals), the essay were in (DIC), and results submitted to ANOVA, Duncan's test (p <0.05). Test one, It was verified that PGPR were tolerant to abiotic stresses of: salinity (80%), temperature (96%) and dry (96%), test two, plants have reached the critical LAS of up to 63% of CC, plants with BRM-32111 had the effect of reducing the damage caused by the water deficit, In biomass 30%, root length 88% in LAS of 30% CC when compared to control plants. Test three, The seeds with BRM-32111 differed from the control in germination, and with BRM 32113 differed from the control in IVG. The plants inoculated with BRM 32111 and BRM32113 had a smaller reduction in root diameter, number of protoxilema pores, cortex thickness, and increase in density of stomata, and increase in carbon assimilation rate (A), efficiency of water use (EUA), carboxylation efficiency rubisco (A / Ci), higher accumulation of chlorophyll a, proline and reduction in the concentration of malonic aldehyde (MDA), in relation to control plants submitted to the same LAS. Test four, the growth of seedlings and plants control rice (CR) was negatively affected by allelopathic compounds. In sowing with residue, plants inoculated with rhizobacteria P. fluorescens BRM-32111 and B. pyrrocinia BRM-32113 induced an increase of 88% in biomass, 3% in the leaf area, 40% and 67% in length and root biomass, respectively, 21% in chlorophyll a, 50% in A, and 63% in the EUA compared to CR control plants CR. These results, evidences that the rhizobacteria are tolerant to the temperature stresses, salinity and osmotic pressure and help to alleviate the harmful effect caused by the water deficit, and increase tolerance of rice plants to stress with allelochemicals in upland rice. / A perda na produtividade de plantas de arroz (Oryza sativa L.) devido aos danos causados por estresses abioticos (déficit hídrico e alelopatia) são recorrentes, o objetico foi avaliar o comportamento de plantas de arroz através das alterações agronômicas e morfológicas sob diferentes lâminas de água no solo (LAS) de 100%, 70 %, 50% e 30 % da capacidade de campo (CC), e identificar as modificações anatômicas e fisiológicos em planta em LAS de 100 % e 50 % da CC induzidas por PGPR (Burkholderia pyrrocinia BRM-32113 e Pseudomonas fluorescens BRM-32111), que indiquem o efeito mitigador dos danos causados pelo déficit hídrico. E em plantios com resíduo de plantas do arroz no solo foi identificar e compreender o efeito da aplicação de rizobactérias em plantas de arroz de terras altas em plantios consecutivos. No primeiro ensaio, as PGPR foram submetidas aos estresses abióticos de temperatura (30, 35 e 40 oC), salinidade (0.5 % e 7.5%) e déficit hídrico (0, -0.2, -0.4, -0.6, -0.8, -1.0 e -1.2 Mpa) simulado por PEG com 10 repetições e avaliado o crescimento. No segundo ensaio, as plantas de arroz inoculadas com: BRM-32111, BRM-32113 e controle, foram submetidas as LAS de: 100, 70, 50 e 30 % da CC, com três vasos por tratamento e cinco plantas por vaso. Foram avaliados: água evapotranspirada e potencial hídrico (ψam), biomassa e comprimento da planta e raiz, área foliar, teor relativo de clorofila. No terceiro ensaio, foram usadas plantas controle, inoculadas com BRM-32111 e BRM-32113, submetidas a 100% e 50% da CC, e mantido até aos 28 dias, e avaliado crescimento, fisiologia e anatomia. No quarto ensaio, foram utilizados quatro tratamentos constituídos de sementes de arroz inoculados com P. fluorescens BRM32111 em solo com resíduo, com B. pyrrocinia BRM-32113 em solo com resíduo e plantas controle em solo com resíduo (CR) e plantas controle em solo sem resíduo (SR) de raízes de plantas de arroz (resíduos de Aleloquímicos), todos os ensaios foram em DIC, e os resultados submetidos ANOVA, teste de Duncan (p < 0.05). No primeiro ensaio, verificou - se que as PGPR foram tolerantes aos estresses abióticos de: salinidade (80%), temperatura (96%) e seca (96%), no segundo ensaio, as plantas atingiram a LAS crítica de até 63 % da CC, as plantas com BRM-32111 tiveram os efeitos amenizador aos danos causado pelo déficit hídrico, na biomassa 30%, comprimento da raiz 88% em LAS de 30% de CC quando comparados a plantas controle. No terceiro ensaio, as sementes com BRM-32111 diferiram do controle na germinação, e com BRM 32113 diferiram do controle no IVG. As plantas inoculadas com BRM 32111 e BRM-32113 tiveram menor redução no diâmetro radicular, número de poros de protoxilema, espessura do córtex, e aumento na densidade de estômatos, e aumento em taxa de assimilação de carbono (A), eficiência do uso da água (EUA) e eficiência de carboxilação da rubisco (A / Ci), maior acumulo de clorofila a, prolina e redução na concentração de aldeído malônico (MDA), em relação a plantas controle submetidas a mesma LAS. No quarto ensaio, o crescimento de plântulas e plantas de arroz controle (CR) foi afetado negativamente pelos compostos alelopáticos. Em semeio com resíduo, plantas inoculadas com as rizobactérias P. fluorescens BRM-32111 e B. pyrrocinia BRM-32113 induziram aumento em 88% na biomassa, 3% na área foliar, 40% e 67% no comprimento e biomassa radicular, respectivamente, 21% na clorofila a, 50% na A, e 63% no EUA comparado as plantas controle CR. Estes resultados, evidenciam que as rizobactérias são tolerantes aos estresses de temperatura, salinidade e pressão osmótica e auxiliam na amenização do efeito danosos causados pelo déficit hídrico, e aumentam a tolerância de plantas de arroz ao estresse com aleloquímicos no arroz de terras altas

<p>Molecular chaperones help many proteins in the cell reach their native conformation. The mechanism with which they do this has been studied extensively, but has not been entirely elucidated. This work is a continuation of the study done by Laila Villebeck et al. (2007) on the conformational rearrangements in the eukaryotic protein actin in interaction with the eukaryotic chaperone TRiC. In this study the intentions were to analyze the protein MreB, a prokaryotic homologue to actin, when interacting with the prokaryotic chaperone GroEL. The purpose was to investigate if the mechanisms of GroEL and TRiC are similar. The analysis of the conformation of MreB was to be made through calculations of fluorescence resonance energy transfer (FRET) between two positions in MreB labeled with fluorescein. A MreB mutant was made through site-specific mutagenesis to enable labeling at a specific position. Another single mutant and a corresponding double mutant needed for these measurements were avaliable from earlier studies. The results from fluorescence measurements on these mutants indicated that the degree of labeling was insufficient for accurate determination of FRET. Suggestions are made on improvements of the experimental approach for future studies.</p>

A major factor in the ability of Pseudomonas fluorescens Pf-5 to act as a biological control agent is its production of antibiotics, including pyoluteorin (PLT), 2,4-diacetylphloroglucinol (2,4-DAPG) and pyrrolnitrin (PRN). The data provided in this thesis demonstrate that the presence of any of these antibiotics in the extracellular milieu affects production of that same antibiotic, as well as others, by Pf-5. Amending the growth medium with antibiotics had multiple effects on secondary metabolism in Pf-5. i) PLT positively regulated its own production, ii) 2,4-DAPG positively regulated its own production. iii) PLT suppressed 2,4-DAPG production. iv) 2,4- DAPG inhibited PLT production. v) PLT suppressed transcription of a heterologous ferric-pyoverdine uptake gene. vi) PRN exerted a slight inhibitory effect on PLT gene transcription and production. PLT autoinduction by Pf-5 was extensively characterized, and was shown to require concentrations of exogenous PLT in the nanomolar range. These low concentrations are comparable to those of many molecules proposed to function in signaling roles. PLT served as a signal between distinct populations of cells within the rhizosphere, where it prompted autoinduction by those cells. Aside from effects of Pf- 5 antibiotics on one another, I also described the positive effect of exogenous PLT on expression of a set of transport genes flanking the PLT biosynthetic gene cluster. Sequence data and experimental evidence suggests that these genes encode a transport apparatus for PLT. The deduced amino acid sequences for four adjacent open reading frames together resemble Type I secretion apparatuses, which typically function in transport of proteins rather than secondary metabolites. The intact transporter genes are necessary for optimal PLT production. Taken together, the data from the studies described herein demonstrate that i) the production of PLT by Pf-5 can affect the production of PLT by neighboring cells, and ii) PLT and other exogenous secondary metabolites have both autoregulatory and cross-regulatory effects in culture. Because Pf-5 derivatives engaged in PLT crossfeeding in the rhizosphere, it is likely that cross-feeding occurs for other secondary metabolites as well. Thus, production of an antibiotic by one cell can profoundly affect secondary metabolism in neighboring cells occupying natural habitats. / Graduation date: 2004

Competitive exclusion has been the mechanism hypothesized to account for the biological control of fire blight disease of pear and apple by the bacterium Pseudomonas fluorescens A506 (A506). Recent laboratory assays demonstrated, however, that A506 produces an antibiotic that is toxic to the fire blight pathogen, Erwinia amylovora, when cultured on media amended with iron (Fe����� or Fe�����). This study investigated this iron-dependent antibiosis by A506 by: 1) examining bioavailability of iron to A506 on blossom surfaces, 2) mutagenizing A506 to disrupt genes involved in antibiotic production, and 3) evaluating suppression of fire blight by A506 when co-treated with an iron chelate (FeEDDHA). Bioavailability of iron on blossoms was investigated with an iron biosensor [iron-regulated promoter (pvd) fused to an ice nucleation reporter gene (inaZ)] in A506. A506 (pvd-inaZ) expressed high ice nucleation activity (INA) on blossoms indicating a low-iron environment unlikely to induce antibiosis by A506. Spraying blossoms with FeEDDHA at concentrations ���0.1 mM significantly suppressed INA by A506 (pvd-inaZ). Transposon mutagenesis was used to generate and select mutants of A506 exhibiting altered antibiotic production profiles. One antibiotic-deficient mutant, A506 Ant���, was recovered; this mutant showed reduced epiphytic fitness on blossoms of apple and pear trees compared to the parent stain, A506. Another mutant, A506 Ant���, lost the characteristic fluorescent phenotype and exhibited iron-independent antibiotic production in defined culture media. A506 Ant��� established high populations on blossoms of apple and pear trees, similar to populations attained by A506, and reduced incidence of fire blight between 20 to 40%, levels comparable to A506 in orchard trials. In orchard trials, A506 was co-treated with FeEDDHA and fire blight suppression was evaluated. Bacterial strains established high populations on blossoms when co-treated with 0.1 mM FeEDDHA or in water. Significantly enhanced suppression of fire blight incidence by antibiotic producing strains of A506 amended with 0.1 mM FeEDDHA was observed in 2 of 5 trials, providing some evidence that iron-induced antibiosis can be a contributing mechanism in disease control. Lack of disease control by the antibiotic deficient strain, A506 GacS, and by 0.1 mM FeEDDHA alone added support to this hypothesis. / Graduation date: 2004

Graduation date: 1998

S'avaluaren 58 soques de Pseudomonas fluorescens i Pantoea agglomerans per la seva eficàcia en el biocontrol de la malaltia causada per l'oomicet Phytophthora cactorum en maduixera i pel nematode formador de gal·les Meloidogyne javanica en el portaempelt GF-677.Es desenvolupà un mètode ex vivo d'inoculació de fulla amb l'objectiu de seleccionar soques bacterianes com a agents de control biològic de P. cactorum en maduixera. Tres soques de P. fluorescens es seleccionaren com a soques eficaces en el biocontrol del patogen en fulles i en la reducció de la malaltia en plantes de maduixera. La combinació de soques semblà millorar la consistència del biocontrol en comparació amb les soques aplicades individualment.Tres soques de P. fluorescens es seleccionaren per la seva eficàcia en la reducció de la infecció de M. javanica en portaempelts GF-677. La combinació d'aquestes soques no incrementà l'eficàcia del biocontrol, però semblà reduir la seva variabilitat. / 58 Pseudomonas fluorescens and Pantoea agglomerans strains were evaluated for their biocontrol efficacy against the oomycete Phytophthora cactorum in strawberry and the root-knot nematode Meloidogyne javanica in GF-677 rootstocks.An ex vivo detached leaf inoculation method was developed to select bacterial strains as biological control agents of P. cactorum in strawberry. Three P. fluorescens strains were selected as effective in biocontrol of the pathogens on leaves and in disease reduction in strawberry plants. Combination of strains improved biocontrol consistency compared to strains applied individually. Three P. fluorescens strains were selected for their efficacy in M. javanica infection reduction in GF-677 rootstocks. Combination of these strains did not increase biocontrol efficacy, but reduced its variability.