F1651, les pili Pap et l’antigène CS31A associé aux antigènes de surface K88 sont tout trois des membres de la famille de type P des facteurs d’adhérence jouant un rôle prépondérant lors de l’établissement d’une maladie causée par des souches Escherichia coli pathogènes, en particulier des souches d’E. coli pathogènes extra-intestinales (ExPEC, Extra-intestinal pathogenic E. coli). Leur expression est sous le contrôle d’un mécanisme de régulation transcriptionnel dépendant de l’état de méthylation de l’ADN, résultant dans l’existence de deux populations définies, l’une exprimant l’adhésine (population ON) et l’autre ne l’exprimant pas (population OFF). Malgré de fortes identités de séquences, ces trois systèmes diffèrent l’un de l’autre, principalement par le pourcentage de cellules ON rencontrées. Ainsi, quand CS31A est systématiquement orienté vers un état considéré comme OFF, F1651 présente une phase ON particulièrement élevée et Pap montre deux états OFF et ON bien distincts, selon le phénotype de départ. La protéine régulatrice sensible à la leucine (Lrp, Leucine-responsive regulatory protein) joue un rôle essentiel dans la réversibilité de ce phénomène épigénétique et il est supposé que les différences de séquences au niveau de la région régulatrice modifient la localisation à ces sites de fixation de Lrp; ce qui résulte, en final, aux différences de phase existant entre CS31A, F1651 et Pap.À l’aide de divers techniques parmi lesquelles l’utilisation de gènes rapporteurs, mutagénèses dirigées et d’analyse des interactions ADN-protéines in vitro, nous montrons dans ce présent projet que la phase OFF prédominante chez CS31A est principalement due à une faible interaction de Lrp avec la région distale de l’opéron clp, et que la présence d’un homologue du régulateur local PapI joue un rôle également clef dans la production de CS31A. Dans le cas de F1651, nous montrons dans cette étude que le taux élevé de cellules en phase ON est dû à une altération dans le maintien de Lrp sur les sites répresseurs 1-3. Ceci est dû à la présence de deux nucléotides spécifiques, situé de part et d’autre du site répresseur 1, qui défavorisent la fixation de Lrp sur ce site précis. Tout comme dans le cas de CS31A, la formation d’un complexe, activateur ou répresseur de la phase ON, dépend également de l’action de du régulatuer local FooI, qui favorise alors le déplacement de Lrp des sites répresseurs 1-3 vers les sites activateurs 4-6. / F1651, the pyelonephritis-associated pili (Pap) and the K88-related surface antigen CS31A are three members of the type P family of adhesive factors that play a key role in the establishment of disease caused by Extra-intestinal Escherichia coli (ExPEC) strains. They are all under the control of methylation-dependent transcriptional regulation that defines the number of fimbriated (ON) and afimbriated (OFF) cells within a clonal population. Despite a high similarity in DNA sequence, these three adhesive systems nonetheless differ in the ratio of ON cells. While CS31A is always turned toward the OFF state, F1651 presents a particularly high level of ON cells and Pap shows two distinct OFF and ON states, depending on the starting phenotype. The leucine-responsive regulatory protein (Lrp) plays an essential role in the reversibility of this epigenetic switch and it is believed that the difference in nucleotides within the regulatory region of each operons could modify the binding of Lrp and, in turn, CS31A, Pap and F1651 phase variation. Using a variety of techniques including gene expression, site-directed mutagenesis, and in vitro protein–DNA interaction analysis, we demonstrate that the preferential OFF state observed in CS31A-positive cells is mainly due to a weak interaction of Lrp with the clp distal region and that the presence of a PapI homologue within the cell plays a key role in CS31A production. For F1651, we show in this study that the high level of ON cells found during F1651 phase variation is due to an altered stability of the DNA complex formed by Lrp at its repressor binding sites 1-3. Again, after each cell cycle, complex formation is modulated by the local regulator FooI (homologue to PapI) which promotes the transit of Lrp toward its activator binding sites 4-6. Furthermore, we identify two nucleotides (T490, G508) surrounding the Lrp-binding site 1 that are critical to maintaining a high OFF to ON switch rate during F1651 phase variation, as well switching Pap fimbriae toward the OFF state.

No description available.

Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2017-02-02T14:03:32Z No. of bitstreams: 1 VictorCarvalhoGalvaoDeFreitas_DISSERT.pdf: 9256729 bytes, checksum: 2b7fb909aa180b8bef8f5de69a706540 (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2017-02-08T20:20:54Z (GMT) No. of bitstreams: 1 VictorCarvalhoGalvaoDeFreitas_DISSERT.pdf: 9256729 bytes, checksum: 2b7fb909aa180b8bef8f5de69a706540 (MD5) / Made available in DSpace on 2017-02-08T20:20:54Z (GMT). No. of bitstreams: 1 VictorCarvalhoGalvaoDeFreitas_DISSERT.pdf: 9256729 bytes, checksum: 2b7fb909aa180b8bef8f5de69a706540 (MD5) Previous issue date: 2016-09-30 / Diversas ferramentas de manuten??o t?m sido empregadas para garantir o correto funcionamento da malha dutovi?ria de transporte de petr?leo, g?s natural e derivados, devido ao importante papel que ela desempenha na cadeia de abastecimento e distribui??o da ind?stria de energia. Entre essas ferramentas, podemos destacar o uso de PIGs (Pipeline Inspection Gauge): dispositivos que viajam no interior dos dutos e s?o capazes de realizar desde uma simples limpeza at? uma inspe??o detalhada de integridade da tubula??o. Neste trabalho, desenvolvemos um sistema de medi??o e supervis?o para um duto de testes cujo principal objetivo ? pesquisar t?cnicas de controle de velocidade para PIGs. Usamos transdutores de press?o ao longo do duto a fim de determinar a posi??o e a velocidade m?dia do PIG. O sistema de supervis?o foi implementado no Elipse SCADA, respons?vel pela visualiza??o e registro de vari?veis do sistema. A interface entre esse sistema e os transdutores foi realizada por meio de um CLP. Os resultados obtidos demonstram que o deslocamento do PIG afeta significativamente o comportamento da press?o no duto e, assim, podem ser usados para estimar a posi??o e a velocidade m?dia do dispositivo por meio dos sensores utilizados. / Several maintenance tools have been employed to ensure the correct operation of the pipeline network to transport oil and gas, due to the important role it plays in the chain of supply and distribution of the energy industry. Among these tools, we can highlight the use of PIG (Pipeline Inspection Gauge) ? a device that travels inside the pipeline and is capable of performing from a simple cleaning to a detailed inspection of pipe integrity. In this work, we developed a measurement and monitoring system for a test pipeline whose main objective is to research PIG speed control techniques. We use pressure transducers along the pipeline in order to determine the position and the average speed of PIG. The supervisory system was implemented in Elipse SCADA, responsible for viewing and recording system variables. The interface between this system and the transducers was done using a Programmable Logic Controller. The results show that displacement of the PIG significantly affects the behavior of the pressure in the pipeline and thus can be used to estimate the position and the average speed of the device by means of the sensors used.

The Regulation No. 1907/2006 on registration, evaluation, authorisation and restriction of chemical substances and on establishment of the European Chemicals Agency, the so-called REACH (Registration, Evaluation, Authorisation and Restriction of Chemical substances) came into effect on 1 June 2007. The purpose of the Regulation has been to ensure safe functioning of the market with chemical substances and preparations and protection of human health and environment against effects of toxic substances. After a complete implementation of the Regulation the only substances and preparations produced and used in the European Union will be those with known toxicological properties, including data about their impact on the health of future generations. The thesis focuses on the description of the REACH Regulation, schedule of its implementation and its potential benefits for the improved effectiveness of rescue and liquidation works performed by members of the Integrated Rescue Service (IZS). Due to very limited sources available at the early stage of REACH implementation, the method selected for the work was consultations with experts directly specializing in the REACH legislation, managers of pharmaceutical or chemical companies managing work with toxic substances and IZS members. Expert opinions have been collected about the legislation process and about the situation in the industry. The Fire Rescue Service uses databases Medis Alarm, Rozex Alarm, TEREX and Trins for planning and implementation of its interventions. In 2013 {--} 2020 I expect bigger quantities of data obtained within the REACH framework to be transmitted into those databases; in that period it will be necessary for the IZS members to become familiar with REACH and with the implemented methods of transmission of toxicological data and to be able to critically evaluate available databases when planning an intervention under conditions of a potential threat posed by toxic substances to the health of IZS members, to the population in the surroundings and to the environment. I recommend to start teaching REACH as a topic at specialized schools so that future graduates are familiar with REACH at least theoretically.

Cytosolic protein degradation is crucial for cellular homeostasis as it orchestrates protein turnover by destruction of misfolded, unstable and abnormal proteins. This process has two main stages: (i) an ATP-dependent stage mediated by unfoldases and proteases, and (ii) an ATP-independent stage mediated by various peptidases. The ATP dependent proteases recognise target proteins and cleave them into smaller peptides. These enzymes comprise the ATPase-family-associated-with-various-cellular-activities domain that is important for unfolding target proteins. Subsequently, unfolded proteins enter a barrel-shaped proteolytic chamber, an architecture conserved throughout prokaryotes, archea and eukaryotes, where the peptide bond is hydrolysed in an ATP-independent manner. The smaller peptides released are broken down by ATP-independent peptidases into free amino acids recycled into the cellular pool. In prokaryotes, major cellular protein degradation functions are performed by Clp and Lon proteases. Earlier studies in our laboratory have shown the role of an ATP-independent peptidase, AminopeptidaseN, in sodium salicylate (NaSal) induced growth inhibition. NaSal belongs to the family of Non-Steroidal Anti-Inflammatory Drugs and its acetyl ester, Aspirin, is a very widely used analgesic. It is produced by plants as a defence response and is known to cause different effects, including xenobiotic stress i. e. stress mediated by compounds which are not naturally produced or expected to be present in the organism in bacteria. In bacteria, salicylate modulates outer membrane proteins, virulence factors, and reduces motility. In addition, NaSal is able induce “phenotypic antibiotic resistance” by binding to MarR and de-repressing the mar operon. NaSal promotes the dissociation of MarR from the promoter site leading to transcription of MarA, a transcription factor that induces several genes that encode the AcrAB-TolC pump which effluxes multiple antibiotics from the cell. The present study investigates the possible roles of ATP-dependent proteases, Lon and Clp, during growth reduction of E. coli induced with high (2-3 mM) amounts of NaSal. The growth of the Lon mutant (lon), but not clpP, was found to be greatly reduced with high doses of NaSal in the media. Our hypothesis was that the lack of Lon led to the accumulation of high amounts of substrate proteins, which led to its greater sensitivity with high doses of NaSal. To identify the substrate protein/s and to better understand the mechanism of action, single and double mutants (with lon) of E. coli lacking several prominent Lon substrates, i.e. MarA, RcsA, StpA, SulA and UmuD, were generated and screened for rescue of growth with 2-3 mM NaSal. MarA, a transcription factor, was identified to be important during NaSal-induced growth reduction. It modulates outer membrane proteins and induces the AcrAB-TolC pump that increases the efflux of antibiotics from the cell. Also, RT-PCR analysis revealed that the levels of marA and its targets, acrA and acrB, were higher in the lon strain suggesting that the MarA protein levels were stabilised the cell in the absence of Lon. Further studies using approriate strains demonstrated that one of the effectors of MarA, i.e. the AcrAB-TolC efflux pump, was not involved in the NaSal-induced growth inhibition of lon. Therefore, in presence of higher doses of NaSal, MarA is upregulated due to de-repression of the operon. The levels of MarA are regulated by Lon via degradation but in the absence of Lon, MarA levels are stabilised and lead to upregulation of MarA and its target genes like AcrAB-TolC. This study identifies higher amounts of MarA to be responsible for NaSal-induced growth inhibition of lon. Subsequently, experiments were conducted to demonstrate the role of MarA and its targets in antibiotic resistance with low dose (0.5 mM) NaSal that does not affect growth. This low dose of NaSal was able to upregulate marA and its target genes, acrA, acrB and tolC. Quantification of antibiotic resistance further revealed an induction in resistance by 0.5 mM NaSal in a MarA- and AcrB-dependent fashion. Studies using atomic force microscopy demonstrated that ciprofloxacin-induced cell elongation was lower in lon due to higher levels of MarA. Therefore, low dose of NaSal is capable of upregulating MarA and inducing antibiotic resistance but does not affect cell growth. This part of the study addresses the roles of Lon protease, its substrate MarA and MarA-induced targets, e.g. AcrB, during NaSal-mediated growth reduction and antibiotic resistance. The factors contributing to antibiotic resistance in bacteria are an important area of study for the global public health care system. Antibiotic resistance can be acquired by transmittance of genetic material, accumulation of antibiotic resistant mutations in the target molecule or can be induced by certain compounds present in the environment like NaSal. For rapid identification of compounds that may behave in a similar fashion as NaSal, a 96-well plate based screen was developed utilising the growth inhibition feature of the lon strain. The compounds were selected on the basis of their structural (phenolic compounds) and functional (Non-Steroidal Anti Inflammatory Drugs or NSAIDs) similarity to NaSal. Through this screen, four compounds were identified that lowered the growth of lon more than that of wild type strain and may be important in inducing phenotypic antibiotic resistance: Acetaminophen (anti-pyretic), Ibuprofen (NSAID), and two phenolic uncouplers, Carbonyl cyanide m-chlorophenyl hydrazone (CCCP) and 2,4-dinitrophenol (2,4-DNP). Notably, another compound Phenylbutazone (NSAID), which is used to lower inflammation in animals, did not reduce the growth of E. coli. RNA expression analysis revealed that these four compounds, but not phenylbutazone, induced the expression of marA and its target gene involved in antibiotic efflux, acrB. Furthermore, dose dependent and comparative studies with Nasal demonstrated differential effects of these four compounds in inducing antibiotic resistance with respect to ciprofloxacin, tetracycline and nalidixic acid. The two uncouplers were much more effective in inducing antibiotic resistance at lower doses than the NSAIDs. As NSAIDs are clinically important compounds, the study suggests that it would be desirable to screen them for induction of antibiotic resistance. The approach elucidated in this study has the potential to identify additional compounds present in the environment that may contribute to antibiotic resistance in bacteria. Overall, this study delineates the roles of Lon protease and its substrate, MarA, during NaSal-mediated responses, involving antibiotic resistance and/or growth reduction in E. coli. In addition, four other compounds were identified that could induce phenotypic antibiotic resistance in E. coli in a MarA-dependent manner. These observations may have implications in the adaptation of bacteria under different environmental conditions.

Syfte:  Syftet med föreliggande översiktsstudie, med fokus på käk- och gomspalt, är att identifiera vilka behandlingsalternativ där tandteknikern inkluderas i LKG (Läpp-, Käk- och Gomspalt)-teamet det finns och vilka av behandlingsalternativen som är att föredra utifrån ansiktsestetik och funktion (äta, andning och tal). Material & metod: Systematiska litteraturstudier granskades utifrån PICO-systemet och sökning i databasen PubMed. Resultat: Endast en artikel funnen i PubMed, uppfyllde urvalskriterierna. Denna artikel inkluderade 20 studier i sin analys, där 3 av dem var mest unika. Från dessa 3 studier kunde en och samma patientens behandling från 2 veckor till 12 år följas. Översiktsstudien gav värdefull information om funktion (tal), estetik, livskvalité och utveckling (längd och språk) samt psykosociala konsekvenser. Analys baserat på de 20 studierna visar att användningen av ortodontisk apparatur inte gett någon större skillnad om apparaturen användes eller ej. Slutsats: Det är oklart vilka behandlingsalternativ som är att föredra för behandling av käk- och gomspalt utifrån ansiktsestetik och funktion. Olika apparaturer kan ge olika resultat, dock kan man inte se någon skillnad på slutresultatet baserat på om en patient använder ortodontisk apparatur eller inte. För att få tydligare svar på frågeställningen behövs ytterligare forskning göras, med flera kliniska uppföljningsstudier som kan resultera i en bättre behandlingsplan för den individuella patienten. / Purpose: The purpose of this overview study, focusing on the cleft of the jaw and palate, is to identify which treatment options that are available where the dental technician is included in the CLP team and which of the treatment options are preferable based on facial aesthetics and function (eating, breathing and speaking). Material & method: An overview was done of systematic reviews based on the PICO-system and searching PubMed database. Results: Only one systematic review was included from PubMed which met the selection criteria. This article included 20 studies, where 3 of them were most unique. From these 3 studies, patient treatment from 2 weeks to 12 years could be followed. This systematic review article provided valuable information on function (speech), aesthetics, quality of life and development (length and language) as well as psychosocial consequences. Analysis based on the 20 studies shows that the use of orthodontic appliances made no major difference whether they were used or not. Conclusion: It is unclear which treatment alternatives that are preferable for the treatment of cleft of the jaw and palate based on facial aesthetics and function.  Different constructions can give different results, however, one cannot see any difference in the treatment outcome based on whether a patient had used an orthodontic appliance or not. Further research including more clinical follow-up studies needs to be done to get clear answers of the research question and with evidence available, resulting in a better treatment plan for the individual patient.

Functionalizable spherical colloidal lignin particles (CLPs) represent a valuable asset for the valorization of lignin side-streams from the pulp industry. The spherical structure allows for the circumvention of the heterogeneous and poorly dispersible structure of the biopolymer. However, organic solvents and alkaline media degrade the particle structure and dissolve the polymers due to their chemical nature and solubility. The solvents will alter the aggregated polymers into irregular shapes that would correspond to inconsistent physicochemical properties. Then, the material will become unusable for advanced material applications, namely wood adhesives. In this study, a replicable process to yield pH ca. 12 stable CLPs for wood adhesives or further functionalization for other advanced material applications was developed and optimized. Lignin was functionalized with cross-linkers, glyoxal or formaldehyde, and selfassembled into spherical structures in the micro emulsification of the organic solution. The formed colloids were partially rotary evaporated to retain organic solvents within the colloidal structures, and then be cured at 73-76 °C until pH stable and further functionalized for advanced material applications. The functionalization with glyoxal was pursued further for its possibly increased reactivity and the health concerns associated with formaldehyde. The process requires the addition of glyoxal to lignin in an acidic organi cmedia at ambient temperature, and the solution to react at 64 °C. Glyoxal is likely added to the polymer structure in its hydrated and dimerized form, and its attachment to lignin should be analyzed through the behavior of glyoxal in different media. The formed colloids were rotary evaporated to an organic solvent content of 60 wt. % of the spheres to allow the occurrence of the curing reaction. These materials were finally cured by thermosetting them at 73-76 °C until pH stable. The particles can be cured with base-catalysis through the controlled addition of the base NaOH(aq). However, the mode and rate of addition of the catalyst are critically important for a nondegradative infusion of a base into solvent present ot removed particles without morphological changes. Further procedural improvement and larger batches are necessary to conduct CLP adhesive experiments. / Funktionaliserbara sfäriska kolloidala ligninpartiklar (CLP) är en värdefull tillgång för valorisering av ligninsidoströmmar från massaindustrin. Den sfäriska strukturen reducerar effekten av den heterogena och dåligt dispergerbara biopolymeren. Organiska lösningsmedel och alkaliska medier försämrar emellertid partikelstrukturen och löser upp polymererna på grund av deras kemiska natur och löslighet. Lösningsmedel kommer att resultera i att de aggregerade polymererna antar oregelbundna former vilket skulle resultera i inkonsistenta fysikalisk-kemiska egenskaper. Därigenom blir materialet oanvändbart för avancerade materialapplikationer, såsom t ex trälim. I denna studie, utvecklades och optimerades en reproducerbar process för att ge pH ca. 12 stabila CLP för trälim eller ytterligare funktionalisering för andra avancerade materialapplikationer. Lignin funktionaliserades med tvärbindare, glyoxal eller formaldehyd och självorganiserades till sfäriskas trukturer genom mikroemulgering av organfasen. De bildade kolloiderna indunstades delvis roterande för att bibehålla det organiska lösningsmedlet i de kolloidala strukturerna och härdades sedan vid 73-76 ° C tills pH-stabilitet och funktionaliserades ytterligare för avancerade materialapplikationer. Funktionaliseringen med glyoxal utfördes också för att reaktiviteteten och begränsa de hälsoproblem som är förknippade med formaldehyd. Förfarandet kräver tillsats av glyoxal till lignin i ett surt organiskt medium vid rumstemperatur för att sedan reageras vid 64 ° C. Glyoxal i dess hydratiserade och dimeriserade form adderas sannolikt till polymeren, och dess kemiska inbindning till lignin kan analyseras genom att undersöka glyoxal uppförande i olika medier. De bildade kolloiderna indunstades till ett organiskt lösningsmedelsinnehåll av 60 viktprocent för att möjliggöra härdning. Dessa material härdades slutligen genom värmehärdning vid 73-76 ° C tills pH var stabilt. Partiklarna kan härdas med baskatalys genom kontrollerad tillsats av basen NaOH (aq). Emellerti där sättet och tillsatshastigheten för katalysatorn kritiskt viktigt. Ytterligare processförbättringar och större satser är nödvändiga för att genomföra CLP-limexperiment.

Sepsis is life-threatening organ dysfunction produced by a dysregulated host response to infection in which neutrophils release neutrophil extracellular traps (NETs). NETs consist of DNA, histones, and antimicrobial peptides which kill pathogens. However, DNA and histones also exert damage by activating the intrinsic pathway of coagulation and inducing endothelial cell death, respectively. AADH, a 15kDa non-anticoagulant unfractionated heparin (UFH), prevents histone-mediated cytotoxicity in vitro and improves survival in septic mice. We explored the effectiveness of Vasoflux, a 5.5kDa low-molecular-weight-heparin as an anti-sepsis treatment as compared to enoxaparin and UFH. Vasoflux has reduced anticoagulant functions and hence reduces the risk of bleeding as compared to enoxaparin or UFH. We showed that UFH, enoxaparin, or Vasoflux at concentrations of up to 13.3uM, 40uM, or 40uM, neutralize histone-mediated cytotoxicity. These results suggest that these glycosaminoglycans (GAGs) are able to neutralize histone-mediated cytotoxicity independent of the AT-binding pentasaccharide. To quantitate the binding affinity between GAGs and histones, surface plasmon resonance was conducted. UFH is a more potent inhibitor of histone-mediated cytotoxicity compared to Vasoflux as UFH has a 10-fold greater binding affinity to histones compared to Vasoflux. To translate our in vitro findings to in vivo, Vasoflux, enoxaparin, and UFH were administered in a murine model of sepsis. Vasoflux at 8mg/kg - 50mg/kg reduced survival and exhibited damage in the lung, liver, and kidney in septic mice compared to 10 mg/kg of UFH or 8mg/kg of enoxaparin. This may be due to Vasoflux and UFH disrupting the DNA-histone complex, thereby releasing free procoagulant DNA. This is evident by our gel electrophoresis experiments, where addition of 1uM Vasoflux or 3.3uM UFH to DNA-histone complexes lead to histone dissociation from DNA. UFH bound to histones may be able to inhibit DNA-mediated thrombin generation, as it retains its anticoagulant properties, demonstrated by UFH-histone complexes attenuating DNA and TF-mediated thrombin generation. In contrast, Vasoflux may not neutralize the procoagulant DNA leading to a hypercoagulable state in the mice. Our study may have important clinical implications as there is an ongoing trial, HALO, which will administer intravenous UFH to patients suspected to have septic shock to reduce mortality. Based on our results, future clinical trials should consider the antithrombin-dependent anticoagulant activity of UFH being used as a sepsis treatment. / Thesis / Master of Science (MSc) / Sepsis is a life threatening condition caused by the body’s extreme response to microbial infection of the blood, whereby neutrophils release traps composed of cell-free DNA (cfDNA), histones, and antimicrobial proteins. In addition to fighting off infections, these traps also exert harmful effects like triggering clotting and killing host cells. Currently, no specific anti-septic drugs exist. Studies have shown that DNase1 (a recombinant protein that digests double stranded cfDNA) or a modified form of heparin (neutralizes histones) improves survival in septic mice. Our goal was to explore the protective effects of Vasoflux, (a non-anticoagulant heparin) and DNase1 in a mouse model of sepsis. We hypothesize that the combined therapy of DNase1 and Vasoflux will improve survival. We found that Vasoflux has minimal blood thinning activity and can prevent histones from killing cells. However, Vasoflux administered into septic mice worsened organ damage and decreased survival. We hypothesize that this damage may be due to Vasoflux’s ability to displace histones from histone-DNA complexes, thereby releasing free DNA, which promotes excessive blood clotting in sepsis.

The first aim of the research was to determine the applicability of certain variables from the Health Belief Model (HBM), the Theory of Reasoned Action (TRA), the risk dimensions from the Psychometric Paradigm, the Common-Sense Model of Illness Representations and the Locus of Control to Italian women’s beliefs and behaviours in relation to screening mammography. These models have predominantly been derived and evaluated with English-speaking persons. The study used quantitative and qualitative methods to enable explanation of research-driven and participant-driven issues. The second aim was to include Italian women in health behaviour research and to contrast the Italian sample with the Anglo-Australian sample to determine if differences exist in relation to their beliefs. In Australia many studies in health behaviour research do not include women whose first language is not English. The third aim was to evaluate the Anti-Cancer Council of Victoria’s (ACCV) Community Language Program (CLP) by: (a) identifying the strengths and weaknesses of the program as seen by the participants; and (b) assessing the impact of the program on women’s knowledge and beliefs about breast cancer, early detection of breast cancer, self-reported and intended breast screening behaviours. The CLP is an information service that uses women’s first language to convey information to women whose first language is not English. The CLP was designed to increase knowledge about breast and cervical cancer. The research used a pre-test-intervention-post-test design with 174 Italian-born and 138 Anglo-Australian women aged 40 years and over. Interviews for the Italian sample were conducted in Italian. The intervention was an information session that related to breast health and screening mammography. Demographic variables were collected in the Pre-Test only. Qualitative open-ended questions that related specifically to the information session were collected in the Post-Test phase of the study. Direct logistic regression was used with the participants’ beliefs and behaviours to identify the relevant variables for language (Italian speaking and English-speaking), attendance to an information session, mammography screening and breast self-examination (BSE) behaviour. Pre- and Post-Test comparisons were conducted using chi-square tests for the non-parametric data and paired sample t-tests for the parametric data. Differences were found between the Italian and Anglo-Australian women in relation to their beliefs about breast cancer screening. The Italian women were: (1) more likely to state that medical experts understood the causes of breast cancer; (2) more likely to feel that they had less control over their personal risk of getting breast cancer; (3) more likely to be upset and frightened by thinking about breast cancer; (4) less likely to perceive breast cancer as serious; (4) more likely to only do what their doctor told them to do; and (5) less likely to agree that there were times when a person has cancer and they don’t know it. A pattern emerged for the Italian and Anglo-Australian women from the logistic regression analyses. The Italian women were much more likely to comply with medical authority and advice. The Anglo-Australian women were more likely to feel that they had some control over their health. Specifically, the risk variable ‘dread’ was more applicable to the Italian women’s behaviour and internal locus of control variable was more relevant to the Anglo-Australian women. The qualitative responses also differed for the two samples. The Italian women’s comments were more general, less specific, and more limited than that of the Anglo-Australian women. The Italian women talked about learning how to do BSE whereas the Anglo-Australian women said that attending the session had reminded them to do BSE more regularly. The key findings and contributions of the present research were numerous. The focus on one cultural group ensured comprehensive analyses, as did the inclusion of an adequate sample size to enable the use of multivariate statistics. Separating the Italian and Anglo-Australian samples in the analyses provided theoretical implications that would have been overlooked if the two groups were combined. The use of both qualitative and quantitative data capitalised on the strengths of both techniques. The inclusion of an Anglo-Australian group highlighted key theoretical findings, differences between the two groups and unique contributions made by both samples during the collection of the qualitative data. The use of a pre-test-intervention-post-test design emphasised the reticence of the Italian sample to participate and talk about breast cancer and confirmed and validated the consistency of the responses across the two interviews for both samples. The inclusion of non-cued responses allowed the researcher to identify the key salient issues relevant to the two groups. The limitations of the present research were the lack of many women who were not screening and reliance on self-report responses, although few differences were observed between the Pre- and Post-Test comparisons. The theoretical contribution of the HBM and the TRA variables was minimal in relation to screening mammography or attendance at the CLP. The applicability of these health behaviour theories may be less relevant for women today as they clearly knew the benefits of and the seriousness of breast cancer screening. The present research identified the applicability of the risk variables to the Italian women and the relevance of the locus of control variables to the Anglo-Australian women. Thus, clear cultural differences occurred between the two groups. The inclusion of the illness representations was advantageous as the responses highlighted ideas and personal theories salient to the women not identified by the HBM. The use of the illness representations and the qualitative responses further confirmed the relevance of the risk variables to the Italian women and the locus of control variables to the Anglo-Australian women. Attendance at the CLP did not influence the women to attend for mammography screening. Behavioural changes did not occur between the Pre- and Post-Test interviews. Small incremental changes as defined by the TTM and the stages of change may have occurred. Key practical implications for the CLP were identified. Improving the recruitment methods to gain a higher proportion of women who do not screen is imperative for the CLP promoters. The majority of the Italian and Anglo-Australian women who attended the information sessions were women who screen. The fact that Italian women do not like talking or thinking about cancer presents a challenge to promoters of the CLP. The key theoretical finding that Italian women dread breast cancer but comply with their doctor provides clear strategies to improve attendance at mammography screening. In addition, the inclusion of lay health advisors may be one way of increasing attendance to the CLP by including Italian women already attending screening and likely to have attended a CLP session. The present research identified the key finding that improving Anglo-Australian attendance at an information session is related to debunking the myth surrounding familial risk of breast cancer and encouraging the Anglo-Australian women to take more control of their health. Improving attendance for Italian women is related to reducing the fear and dread of breast cancer and building on the compliance pattern with medical authority. Therefore, providing an information session in the target language is insufficient to attract non-screeners to the session and then to screen for breast cancer. Suggestions for future research in relation to screening mammography were to include variables from more than one theory or model, namely the risk, locus of control and illness representations. The inclusion of non-cued responses to identify salient beliefs is advantageous. In addition, it is imperative to describe the profile of the cultural sample in detail, include detailed descriptions of the translation process and be aware of the tendency of Italian women to acquiesce with medical authority.

<p>Unlike in the greenhouse, plants need to cope with many environmental stresses under natural conditions. Among these conditions are drought, waterlogging, excessive or too little light, high or low temperatures, UV irradiation, high soil salinity, and nutrient deficiency. These stress factors can affect many biological processes, and severely retard the growth and development of higher plants, resulting in massive losses of crop yield and wood production. Plants have developed many protective mechanisms to survive and acclimate to stresses, such as the rapid induction of specific molecular chaperones and proteases at the molecular level. Molecular chaperones mediate the correct folding and assembly of polypeptides, as well as repair damaged protein structures caused by stress, while proteases remove otherwise non-functional and potentially cytotoxic proteins. </p><p>The Clp/Hsp100 family is a new group of chaperones that consists of both constitutive and stress-inducible members. Besides being important chaperones, many Clp/Hsp100 also participate in protein degradation by associating with the proteolytic subunit ClpP to form the Clp protease complex. Higher plants have the greatest number and complexity of Clp proteins than any other group of organisms, and more than 20 different Clp isomers in plants have been identified (Paper I). Because of this diversity, we have adopted a functional genomics approach to characterise all Clp proteins in the model plant Arabidopsis thaliana. Our ongoing research strategy combines genetic, biochemical and molecular approaches. Central to these has been the preparation of transgenic lines for each of the chloroplast Clp isomers. These transgenic lines will be analysed to understand the function and regulation of each chloroplast Clp protein for plant growth and development.</p><p>In Paper II, an Arabidopsis thaliana cDNA was isolated that encodes a homologue of bacterial ClpX. Specific polyclonal antibodies were made and used to localise the ClpX homologue to plant mitochondria, consistent with that predicted by computer analysis of the putative transit peptide. In addition to ClpX, a nuclear-encoded ClpP protein, termed ClpP2, was identified from the numerous ClpP isomers in Arabidopsis and was also located in mitochondria. Relatively unchanged levels of transcripts for both clpX and clpP2 genes were detected in various tissues and under different growth conditions. Using β-casein as a substrate, plant mitochondria possessed an ATP-stimulated, serine-type proteolytic activity that could be strongly inhibited by antibodies specific for ClpX or ClpP2, suggesting an active ClpXP protease.</p><p>In Paper III, four nuclear-encoded Clp isomers were identified in Arabidopsis thaliana: ClpC1 and ClpP3-5. All four proteins are localized within the stroma of chloroplasts, along with the previously identified ClpD, ClpP1 and ClpP6 proteins. Potential differential regulation among these Clp proteins was analysed at both the mRNA and protein level. A comparison between different tissues showed increasing amounts of all plastid Clp proteins from roots to stems to leaves. The increases in protein were mirrored at the mRNA level for most ClpP isomers but not for ClpC1, ClpC2 and ClpD and ClpP5, which exhibited little change in transcript levels. Potential stress induction was also tested for all chloroplast Clp proteins by a series of brief and prolonged stress conditions. The results reveal that these proteins, rather than being rapidly induced stress proteins, are primarily constitutive proteins that may also be involved in plant acclimation to different physiological conditions. </p><p>In Paper IV, antisense repression transgenic lines of clpP4 were prepared and then later characterised. Within the various lines screened, up to 90% of ClpP4 protein content was specifically repressed, which also led to the down-regulation of ClpP3 and ClpP5 protein contents. The repression of clpP4 mRNA retarded the development of chloroplasts and the differentiation of leaf mesophyll cells, resulting in chlorotic phenotypes. The chlorosis was more severe in young than in mature leaves due likely to the developmental expression pattern of the ClpP4 protein. Chlorotic plants eventually turned green upon aging, accompanied by a recovery in the amount of the ClpP4 protein. The greening process could be affected by the light quantity, either by altering the photoperiod or light intensity.</p>