The use of fluorescent probes to assess viability of the plant pathogenic bacterium Clavibacter michiganensis subsp. michiganensis by flow cytometry

Determination of the viability of bacteria by the conventional plating technique is a time-consuming process. Methods based on enzyme activity or membrane integrity are much faster and may be good alternatives. Assessment of the viability of suspensions of the plant pathogenic bacterium Clavibacter michiganensis subsp. michiganensis (Cmm) using the fluorescent probes Calcein acetoxy methyl ester (Calcein AM), carboxyfluorescein diacetate (cFDA), and propidium iodide (PI) in combination with flow cytometry was evaluated. Heat-treated and viable (non-treated) Cmm cells labeled with Calcein AM, cFDA, PI, or combinations of Calcein AM and cFDA with PI, could be distinguished based on their fluorescence intensity in flow cytometry analysis. Non-treated cells showed relatively high green fluorescence levels due to staining with either Calcein AM or cFDA, whereas damaged cells (heat-treated) showed high red fluorescence levels due to staining with PI. Flow cytometry also allowed a rapid quantification of viable Cmm cells labeled with Calcein AM or cFDA and heat-treated cells labeled with PI. Therefore, the application of flow cytometry in combination with fluorescent probes appears to be a promising technique for assessing viability of Cmm cells when cells are labeled with Calcein AM or the combination of Calcein AM with PI.

Synthesis, structural and morphological characterization of CeO2-ZnO nanosized powder systems from Pechini's method

This work reports on the investigation of nanosized CeO2-ZnO systems prepared by Pechini's method. The structural and morphological characterization of CeO2-ZnO systems as well as the characterization of CeO2 and ZnO separately, showed that the employed method result in powders with spheroidal particles whose size are in the range 30 - 200 nm, which is appropriate to provide homogeneous suspensions. The ZnO present in the prepared mixed oxides seems to increase particle size distribution and to influence the arrangement of the particles after powder dispersion.

Synthesis and characterization of Fe(III)-doped ceramic membranes of titanium dioxide and its application in photoelectrocatalysis of a textile dye

Pure and Fe(III)-doped TiO2 suspensions were prepared by the sol gel method with the use of titanium isopropoxide (Ti(OPri)4) as precursor material. The properties of doped materials were compared to TiO2 properties based on the characterization by thermal analysis (TG-DTA and DSC), X-ray powder diffractometry and spectroscopy measurements (FTIR). Both undoped and doped TiO2 suspensions were used to coat metallic substrate as a mean to make thin-film electrodes. Thermal treatment of the precursors at 400ºC for 2 h in air resulted in the formation of nanocrystalline anatase TiO2. The thin-film electrodes were tested with respect to their photocatalytic performance for degradation of a textile dye in aqueous solution. The plain TiO2 remains as the best catalyst at the conditions used in this report.

Cotton resistance to ramulose and variability of Colletotrichum gossypii f. sp. cephalosporioides

Four cultivars and 21 lines of cotton were evaluated for resistance to ramulose (Colletotrichum gossypii f. sp. cephalosporioides) in a field where the disease is endemic. The seeds of each genotype were planted in 5 x 5 m plots with three replications. The lines CNPA 94-101 and 'CNPA Precoce 2'were used as standard susceptible and resistant references, respectively. The disease incidence (DI) was calculated from the proportion of diseased plants in the plot. The disease index (DIn) was calculated from the disease severity using a 1 to 9 scale, and was evaluated at weekly intervals starting 107 days after emergence. The data collected was used to calculate the area under disease progress curve (AUDPC). In general, the DIn increased linearly with time and varied from 20.0 to 57.1 and AUDPC from 567 to 1627 among the genotypes which could be clustered in to two distinct groups. The susceptible group contained two cultivars and nine lines and the resistant group contained one cultivar and 12 lines. The relationship between disease index and evaluation times was linear for the 25 genotypes tested. The line CNPA 94-101, used as susceptible standard, was the most susceptible with an average DI = 83.4, DIn = 57.1 and AUDPC = 1627.7. The line CNPA 96-08 with DI = 37.8, DIn = 20.0 and AUDPC = 567.7 was the most resistant one. Among the commercial cultivars 'IAC 22' was the most susceptible and 'CNPA Precoce 2', used as resistant standard was the most resistant. The variability in virulence of the pathogen was studied by spray inoculating nine genotypes with conidial suspensions (10(5)/mL) of either of the 10 isolates. The disease severity was evaluated 30 days later using a scale of 1 to 5. The virulence of the isolate was expressed by DIn. All the isolates were highly virulent but their virulence avaried for several genotypes and could be clustered in two distinct groups of less and more virulent isolates. The isolate MTRM 14 from Mato Grosso was the least virulent while Minas Gerais was the most virulent, with DIn of 6.36 and 46.47, respectively. In this experiment the line HR 102 and the cultivar 'Antares' were the most resistant ones with DIns of 18.32 and 19.14, respectively.

Happidispersion karakterisointi laboratoriosekoittimessa

Työn tarkoituksena oli tarkastella uutta kuvantamistekniikkaa käyttäen happikaasun dispergointia keskisakeuksisen massasuspension joukkoon laboratoriosekoittimessa. Työssä pyrittiin tarkastelemaan muodostuvan dispersion homogeenisuutta neljästä eri kuvauspisteestä sekoittimen kannesta ja kyljestä. Samalla tarkasteltiin myös sekoittimen tehonkulutusta sekä tehonkulutuksen ja aikaansaadun dispersion välistä yhteyttä. Työn yhtenä tarkoituksena oli myös tarkastella uuden kuvantamistekniikan mahdollisuuksia tämäntyyppisissä sovellutuksissa, sillä työ kuuluu PulpVision-projektiin, jossa kehitetään massa- ja paperiteollisuuden uusia konenäkösovellutuksia. Työn kokeellinen osuus koostui sekoituskokeista, joissa tarkasteltiin neljästä kuvauspisteestä kahdella sekoittimen nopeudella mänty- ja koivususpensioihin muodostuvaa kuplakokojakaumaa. Sekoituskokeiden lisäksi tehtiin tehonkulutuskokeita, joissa tarkasteltiin sekoittimen tehonkulutusta sekoittimen täyttöasteen funktiona koivu- ja mäntysuspensioilla sekä vedellä. Työn tuloksien perusteella todettiin, että koivususpensiosta havaittujen kuplien pinta-ala oli noin puolet mäntysuspensiosta havaittujen kuplien pinta-alasta. Sekoittimen roottorin pyörimisnopeuden puolittuessa suspensioon dispergoidun hapen kuplakoko kasvoi huomattavasti. Neljästä kuvausyhteestä tarkasteltuna havaittiin pienimpien kuplien esiintyvän sekoittimen alaosassa. Mäntysuspension tehonkulutuksen havaittiin kasvavan viidenneksellä, kun sekoittimen täyttöaste kasvoi 10 %, kun taas koivususpension tehonkulutuksen kasvu oli tästä vain puolet. Kuvantamislaitteiston todettiin olevan tämänkaltaiseen sovellutukseen riittävä, varsinkin kun valonlähteenä käytetään pulssilaseria.

Kaasun dispergoituminen tehdasmittakaavaisessa keskisakean massan sekoittimessa

Työn tarkoituksena oli selvittää Proto 10 kemikaalisekoittimen kyky dispergoida kaasua keskisakean valkaistun mäntysellun joukkoon. Työssä käytettiin kuvantamistekniikkaa, jonka avulla pystyttiin näkemään sekoittimen ja putkiston sisälle sekoitustapahtuman aikana. Muodostunutta dispersiota tarkasteltiin kolmesta kuvauspisteestä, joista yksi sijaitsi sekoittimen pesässä ja kaksi putkistossa sekoittimen jälkeen. Työssä verrattiin myös laboratoriosekoittimella saatuja tuloksia teollisen mittakaavan sekoittimella saatuihin tuloksiin, sekä määritettiin tarvittava pinta-aktiivisen aineen konsentraatio, jolla saavutettiin ruskeaa massaa vastaava vaahtoaminen. Työn kokeellinen osuus koostui kolmesta osasta. Ensimmäisessä vaiheessa tutkittiin ruskeasta mänty- ja koivumassasta lingotun suodoksen vaahtoamista ja verrattiin sitä vedellä ja pesuaineella saatavaan vaahtoon. Toisessa vaiheessa suoritettiin referenssiajot laboratoriosekoittimen ja teollisen mittakaavan sekoittimen vertailua varten Quantum Mark IV laboratoriosekoittimella. Kolmannessa vaiheessa tutkittiin Proto 10 sekoittimen kykyä dispergoida kaasua mäntysellun joukkoon eri pyörimisnopeuksilla ja virtaamilla. Työn tuloksien perusteella energiankäytön kannalta paras pyörimisnopeus Proto 10 sekoittimelle on 1500 min-1. Nostamalla kierrosnopeutta yli tämän ei saavutettu merkittävää parannusta dispersion laadussa varsinkaan suuremmilla virtaamilla. Virtaamalla todettiin olevan suuri merkitys sekoitustulokseen. Laboratoriosekoittimella tehtyjen kokeiden todettiin vastaavan parhaiten teollisen mittakaavan tuloksia pyörimisnopeudella 1200 min-1.

Determination of slurry’s viscosity using case based reasoning approach

Case-based reasoning (CBR) is a recent approach to problem solving and learning that has got a lot of attention over the last years. In this work, the CBR methodology is used to reduce the time and amount of resources spent on carry out experiments to determine the viscosity of the new slurry. The aim of this work is: to develop a CBR system to support the decision making process about the type of slurries behavior, to collect a sufficient volume of qualitative data for case base, and to calculate the viscosity of the Newtonian slurries. Firstly in this paper, the literature review about the types of fluid flow, Newtonian and non-Newtonian slurries is presented. Some physical properties of the suspensions are also considered. The second part of the literature review provides an overview of the case-based reasoning field. Different models and stages of CBR cycles, benefits and disadvantages of this methodology are considered subsequently. Brief review of the CBS tools is also given in this work. Finally, some results of work and opportunities for system modernization are presented. To develop a decision support system for slurry viscosity determination, software application MS Office Excel was used. Designed system consists of three parts: workspace, the case base, and section for calculating the viscosity of Newtonian slurries. First and second sections are supposed to work with Newtonian and Bingham fluids. In the last section, apparent viscosity can be calculated for Newtonian slurries.

Studies in offset ink setting

The offset printing process is complex and involves the meeting of two essentially complex materials, printing ink and paper, upon which the final product is formed. It can therefore be expected that a multitude of chemical and physical interactions and mechanisms take place at the ink-paper interface. Interactions between ink and paper are of interest to both the papermakers and ink producers, as they wish to achieve better quality in the final product. The objective of this work is to clarify the combined influence of paper coating structure, printing ink and fountain solution on ink setting and the problems related to ink setting. A further aim is to identify the mechanisms that influence ink setting problems, and to be able to counteract them by changing properties of the coating layer or by changing the properties of the ink. The work carried out for this thesis included use of many techniques ranging from standard paper and printability tests to advanced optical techniques for detection of ink filaments during ink levelling. Modern imaging methods were applied for assessment of ink filament remain sizes and distribution of ink components inside pigment coating layers. Gravimetric filtration method and assessment of print rub using Ink-Surface-Interaction-Tester (ISIT) were utilized to study the influence of ink properties on ink setting. The chemical interactions were observed with the help of modified thin layer chromatography and contact angle measurements using both conventional and high speed imaging. The results of the papers in this thesis link the press operational parameters to filament sizes and show the influence of these parameters to filament size distribution. The relative importance between the press operation parameters was shown to vary. The size distribution of filaments is important in predicting the ink setting behaviour, which was highlighted by the dynamic gloss and ink setting studies. Prediction of ink setting behaviour was also further improved by use of separate permeability factors for different ink types in connection to filtration equations. The roles of ink components were studied in connection to ink absorption and mechanism of print rub. Total solids content and ratio of linseed oil to mineral oil were found to determine the degree of print rub on coated papers. Wax addition improved print rub resistance, but would not decrease print rub as much as lowering the total solids content in the ink. Linseed oil was shown to absorb into pigment coating pores by mechanism of adsorption to pore walls, which highlights the need for sufficient pore surface area for improved chromatographic separation of ink components. These results should help press operators, suppliers of printing presses, papermakers and suppliers to papermakers, to better understand the material and operating conditions of the press as it relates to various print quality issues. Even though paper is in competition with electronic media, high quality printed products are still in demand. The results should provide useful information for this segment of the industry.

Rheological behavior of mixed drink of annona and milk

In this study the rheological behavior in different temperatures (0; 6; 18 e 24 ºC) and physicochemical parameters of integral annona (Annona squamosa) pulp and the annona pulp with milk in different percentages pulp/milk (75g of annona pulp/25g of milk, 50g of annona pulp/50g of milk, 25g of annona pulp/75g of milk) have been availed, in order to verify the effect of temperature and pulp concentration in the rheological behavior of these beverages. To obtain the rheological parameters a concentric cylinder rheometer has been used and the rheograms were analyzed using the Ostwald-de-Wael (power Law) and Herschel-Bulkley models. The physicochemical parameters (sugars, pH, ash, acidity and soluble solids) were determined in order to establish correlations with the rheological behavior. Finally, the best results had been obtained using the Herschel-Bulkley model; the low values for the behavior index (n <1) obtained confirm the pseudoplastic behavior of all samples.

Verhopäällystyspastojen analyysimenetelmät

Diplomityön tarkoitus oli selvittää verhopäällystyspastoille sopivia analyysimenetelmiä. Verhopäällystyksessä onnistunut päällystystapahtuma vaatii venymäviskositeetin ja pintajännityksen hyvää hallintaa. Kirjallisuusosassa käsiteltiin verhopäällystystä, verhopäällystyspastojen koostumusta, reologiaa ja pintajännitystä. Kirjallisuusosassa käsiteltiin lisäksi verhopäällystyspastojen reologian ja pintajännityksen mittaamiseen soveltuvia mittausmenetelmiä. Verhopäällystyksen luonteen vuoksi kirjallisuusosassa syvennyttiin venymäviskositeetin ja dynaamisen pintajännityksen mittaamiseen tarkoitettuihin menetelmiin. Kokeellisessa osassa tutkittiin päällystyspastasarjojen reologiaa ja pintajännitystä verhopäällystystä varten. Osaan päällystyspastoista luotiin venymäviskositeettia ja osasta laskettiin pintajännitystä. Venymäviskositeetin mittaamista varten työssä käytettiin ACAV A2 -reometriin liitettyjä teräsreikälevyjä. Dynaamisen pintajännityksen mittaamista varten työssä käytettiin KSV BPA-800P -pintajännitysmittaria. ACAV A2 -reometriin liitettyjen teräsreikälevyjen (reiän sisähalkaisija 0,5 tai 0,7 mm) avulla mitattiin venymäviskositeettia kuvaavia Eulerin lukuarvoja onnistuneesti suurilla kiintoainepitoisuuksilla (50, 60 tai 65 p %). Erikoispaksuntajan määrää lisäämällä onnistuttiin luomaan huomattavaa venymäviskositeettia. Kiintoainepitoisuuden kasvaessa kasvoi myös venymäviskositeetti. Tavanomaisille paksuntajille mitattiin hieman kohonneita venymäviskositeetteja verrattuna referenssipäällystyspastaan. Pigmenttikoostumuksella (kalsiumkarbonaatti/kaoliini) ei näyttänyt olevan vaikutusta venymäviskositeettiin, tai vaikutus oli suhteellisen pieni. Dynaamisen pintajännityksen mittaamista varten käytössä ollut KSV BPA-800P -pintajännitysmittari ei toiminut luotettavasti, vaikka näytteitä laimennettiin. Kiintoainepitoisuudessa 10 p-% olleilla laimennoksilla saavutettiin analysoinnin kannalta parhaat tulokset. Tuloksista saatiin kuitenkin viitteitä, että kyseinen mittari voisi olla potentiaalinen menetelmä dynaamisen pintajännityksen mittaamiseksi.

Evaluation of Mycobacterium avium subsp. paratuberculosis faecal culture protocols and media

Paratuberculosis is an important enteritis of ruminants caused by Mycobacterium avium subsp. paratuberculosis (Map). The disease is officially considered exotic in Brazil, but recent serological surveys and the isolation of the agent suggest it may occur in our herds. The aim of this study was to evaluate three different formulations of Herrold's egg yolk agar with mycobactin J (HEYM) and four faecal culture protocols considering their ability for Map growth as well as cost and ease of application. Three formulations of HEYM were inoculated with two suspensions of Map. Spiked faeces and naturally contaminated faecal samples were treated by the four faecal culture protocols. Centrifugation protocol and HEYM recommended by OIE showed the best results on the recovery of Map.

Influence of reactant absorption and dissolution in heterogeneous precipitation processes

In this research work, the aim was to investigate the volumetric mass transfer coefficient [kLa] of oxygen in stirred tank in the presence of solid particle experimentally. The kLa correlations as a function of propeller rotation speed and flow rate of gas feed were studied. The O2 and CO2 absorption in water and in solid-liquid suspensions and heterogeneous precipitation of MgCO3 were thoroughly examined. The absorption experiments of oxygen were conducted in various systems like pure water and in aqueous suspensions of quartz and calcium carbonate particles. Secondly, the precipitation kinetics of magnesium carbonate was also investigated. The experiments were performed to study the reactive crystallization with magnesium hydroxide slurry and carbon dioxide gas by varying the feed rates of carbon dioxide and rotation speeds of mixer. The results of absorption and precipitation are evaluated by titration, total carbon (TC analysis), and ionic chromatrography (IC). For calcium carbonate, the particle concentration was varied from 17.4 g to 2382 g with two size fractions: 5 µm and 45-63 µm sieves. The kLa and P/V values of 17.4 g CaCO3 with particle size of 5µm and 45-63 µm were 0.016 s-1 and 2400 W/m3. At 69.9 g concentration of CaCO3, the achieved kLa is 0.014 s-1 with particle size of 5 µm and 0.017 s-1 with particle size of 45 to 63 µm. Further increase in concentration of calcium carbonate, i.e. 870g and 2382g , does not affect volumetric mass transfer coeffienct of oxygen. It could be concluded from absorption results that maximum value of kLa is 0.016 s-1. Also particle size and concentration does affect the transfer rate to some extend. For precipitation experiments, the constant concentration of Mg(OH)2 was 100 g and the rotation speed varied from 560 to 750 rpm, whereas the used feed rates of CO2 were 1 and 9 L/min. At 560 rpm and feed rate of CO2 is 1 L/min, the maximum value of Mg ion and TC were 0.25 mol/litre and 0.12 mol/litre with the residence time of 40 min. When flow rate of CO2 increased to 9 L/min with same 560 rpm, the achieved value of Mg and TC were 0.3 mol/litre and 0.12 mol/L with shorter residence time of 30 min. It is concluded that feed rate of CO2 is dominant in precipitation experiments and it has a key role in dissociation and reaction of magnesium hydroxide in precipitation of magnesium carbonate.

Slurry flows in progressive cavity and in hose pumps

The aim of this thesis was to research how slurry’s viscosity and rheology affect to pumping in peristaltic hose pump and in eccentric progressive cavity pump. In addition, it was researched the formed pressure pulsation in hose pump. Pressure pulsation was studied by pumping different slurries and by using different pipe materials. Pressure and power curves were determined for both used pumps. It was also determined NPSHR curve for the progressive cavity pump. Literature part of the thesis considered to distribute fluids to different rheology types, as well as theories and models to identify different rheology types. Special attention was paid to non-Newtonian fluids, which were also used in experimental part of this thesis. In addition, the literature part discusses about pumps, parameters for pump sizing, and pressure pulsation in hose pump. Starch, bentonite, and carboxymethyl cellulose slurries were used in the experimental part of this thesis. The slurries were pumped with Flowrox peristaltic hose pump (LPP-T32) and eccentric progressive cavity pump (C10/10). From the each slurry was taken a sample, and the samples were analyzed for concentration, viscosity and rheology type. The used pipe materials in pressure pulsation experiments were steel and elastic, and it was also used a prototype of pulsation dampener. The pulsation experiments indicated that the elastic pipe and the prototype of pulsation dampener attenuated pressure pulsation better than the steel pipe at low pressure levels. The differences between different materials disappeared when pressure level and pump rotation speed increased. In slurry experiments, pulsation was different depending on rheology and viscosity of the slurry. According to experiments, the rheology did not significantly affect to pump power consumption or efficiency.

Interphase cytogenetics using fluorescence in situ hybridization: an overview of its application to diffuse and solid tissue

Interphase cytogenetics, utilizing fluorescence in situ hybridization (FISH) techniques, has been successfully applied to diffuse and solid tissue specimens. Most studies have been performed on isolated cells, such as blood or bone marrow cells; a few have been performed on cells from body fluids, such as amniotic fluid, urine, sperm, and sputum. Mechanically or chemically disaggregated cells from solid tissues have also been used as single cell suspensions for FISH. Additionally, intact organized tissue samples represented by touch preparations or thin tissue sections have been used, especially in cancer studies. Advantages and pitfalls of application of FISH methodology to each type of specimen and some significant biological findings achieved are illustrated in this overview.

E. coli a-hemolysin: a membrane-active protein toxin

Alpha-Hemolysin is synthesized as a 1024-amino acid polypeptide, then intracellularly activated by specific fatty acylation. A second activation step takes place in the extracellular medium through binding of Ca2+ ions. Even in the absence of fatty acids and Ca2+ HlyA is an amphipathic protein, with a tendency to self-aggregation. However, Ca2+-binding appears to expose hydrophobic patches on the protein surface, facilitating both self-aggregation and irreversible insertion into membranes. The protein may somehow bind membranes in the absence of divalent cations, but only when Ca2+ (or Sr2+, or Ba2+) is bound to the toxin in aqueous suspensions, i.e., prior to its interaction with bilayers, can a-hemolysin bind irreversibly model or cell membranes in such a way that the integrity of the membrane barrier is lost, and cell or vesicle leakage ensues. Leakage is not due to the formation of proteinaceous pores, but rather to the transient disruption of the bilayer, due to the protein insertion into the outer membrane monolayer, and subsequent perturbations in the bilayer lateral tension. Protein or glycoprotein receptors for a-hemolysin may exist on the cell surface, but the toxin is also active on pure lipid bilayers.