Highly functional binding surface for miniaturised solid-phase immunoassays. - A spot story -

Several bioaffinity assays are based on the detection of an analyte which is bound on a solid substrate via biochemical interaction. These so called solid phase assays are based on the adhesion of the primary binding partner on a solid surface, which then binds the analyte to be detected. In this thesis work a novel solid phase based assay technology, known as spot technology, was developed. The spot technology is based on combination of high-capacity solid phases, concentrated in a spot format, utilising modified streptavidin molecules and recombinant antibody fragments. The reduction of the solid phase binding surface to a size of a spot enabled denser binding of the target molecules, providing improved signal intensities and signal-to-background ratio when applied in different solid phase immunoassays. Streptavidin-biotin interactions are commonly utilised in numerous different bioaffinity assays and the ultimate nature of streptavidin to bind biotin is among the strongest non-covalent interaction reported between two biomolecules. In this study native core streptavidin was chemically modified to provide polymerised streptavidin molecules with altered adsorption properties. These streptavidin conjugates, when coated onto polystyrene surface, provided enhanced biotin binding capacity and surface stability when compared to a reference coating constructed with native streptavidin. Furthermore, the combination of chemically modified streptavidin, sitespecifically biotinylated antibody fragments and the spot coating technology provided highly dense solid phase coating with improved binding properties. The performance of the spot assay technology was further demonstrated in different immunoassay configurations. Human thyroid stimulating hormone (TSH) and human cardiac troponin I (cTnI) were used as model analytes to show the applicability of the highly sensitive spot-based solid-phase immunoassay for detection of very low levels of analytes. It was demonstrated that the spot technology provided an assay concept with enhanced sensitivity and short turn-around times, characteristics that are highly suitable for point-of-care applications.

Prognosis of after spot volumes at the Scandinavian electricity market

The Thesis is dedicated to development of an operative tool to support decision making in after spot trading on the Nordic electricity market. The basics of the Nordic electricity market, trading mechanisms on the spot and after spot markets are presented in the Thesis. Mathematical equations that describe electricity balance condition in the power system are offered. The main driving factors that impact deviation of actual electricity balance from the scheduled one (object) in the power system have been explored and mathematically defined. The behavioral model of the object and principal trends in change of state of the object under an impact of the driving factors are determined with the help of regression analysis made in Microsoft Office Excel. The behavioral model gives an indication for the total regulation volume (Elbas trades volume, volume of regulation market, balance power) for a certain hour that serves as the base input in estimating prices on the after spot markets. Proposals for development of methodologies of forecasting the after spot electricity prices are offered.

Anàlisi d'un spot televisiu segons el model proposat per Frederic Chaume : Estudi d'un cas: Beauty is nothing without brains

L’estudi de les situacions comunicatives sempre m’ha atret de manera significativa. El món de la publicitat, també. L’assignatura de Semiòtica audiovisual ha estat el vincle que ha inspirat el tema d’aquest treball. L’idea inicial era la d’analitzar des d’una perspectiva semiòtica un document audiovisual, concretament, Beauty is nothing without brains, un espot televisiu de Mercedes-Benz emès als Estats Units l’any 2007. Es pretenia identificar i analitzar cadascun dels elements que configuren la comunicació audiovisual i presentar possibles versions traductològiques del mateix. La dificultat de crear un esquema d’anàlisi que em permetés estructurar-ne l’estudi va conduir-me a Frederic Chaume. És llavors quan aconsegueixo concretar el tema del treball: es pretén analitzar aquest document audiovisual seguint el model proposat per Chaume, responent a un hipotètic encàrrec de traducció el qual demana la seva adaptació al mercat català.

Desenvolvimento de um spot test para o monitoramento da atividade da peroxidase em um procedimento de purificação

In this work we describe both a chromatographic purification procedure and a spot test for the enzyme peroxidase (POD: EC 1.11.1.7). The enzyme was obtained from crude extracts of sweet potatoes and the chromatographic enzyme purification procedure resulted in several fractions. Therefore a simple, fast and economic spot test for monitoring peroxidase during the purification procedure was developed. The spot test is based on the reaction of hydrogen peroxide and guaiacol, which is catalyzed by the presence of peroxidase yielding the colored tetraguaiacol.

Epidemiology of apple leaf spot

Apple leaf spot (ALS) caused by Colletotrichum spp. is a major disease of apple (Malus domestica) in Southern Brazil. The epidemiology of this disease was studied in experiments carried out in the counties of Passo Fundo and Vacaria, State of Rio Grande do Sul, from February 1998 to October 2000. The disease was found in all the six apple orchards sampled in the growing seasons of 1997/98 and 1998/99. The fungus isolates associated with ALS fit the characteristics of C. gloeosporioides (75%), C. acutatum (8%), and Colletotrichum sp. (17%). The pathogen overwintered in dormant buds and twigs but not in dropped leaves or fruit mummies. Two sprays of copper oxychloride (at 0.3%) reduced the fungus initial inoculum by 65-84.6% in buds and 85.6-93.7% in twigs, but had no effect on the early season progress of the disease. Disease severity increased proportionally to elevation of temperature from 14 to 26-28 °C. At 34 °C, however, infection was completely inhibited. The duration of leaf wetness required for infection ranged from two hours at 30 °C to 32 h at 16 °C. The relationship of temperature (T) and leaf wetness (W) to disease severity (Y) was represented by the model equation Y = 0.00145[((T-13)1.78)((34.01-T )1.09)] * 25/[1+14 exp(-0.137W)], R² = 0.73 and P < 0.0001. Currently, this information is being used to manage the disease and to validate a forecast system for ALS.

Caracterização do Tomato chlorotic spot virus isolado de jiló no Vale do Paraíba, Estado de São Paulo

Os tospovírus são responsáveis por perdas significativas em diversas culturas, principalmente solanáceas. No município de São José dos Campos (SP), plantas de jiló (Solanum gilo) apresentando sintomas de mosaico, bolhosidades, nanismo e queda acentuada da produção foram coletadas para análise. Visando a caracterização do agente causador dos sintomas, testes biológicos, elétrono microscópicos, sorológicos e moleculares foram realizados. Através de inoculação mecânica em plantas indicadoras das famílias Amaranthaceae, Chenopodiaceae e Solanaceae obtiveram-se resultados típicos aos esperados para tospovírus. Ao microscópio eletrônico de transmissão, observaram-se, em contrastação negativa, partículas pleomórficas com diâmetro entre 80 e 110 nm e em cortes ultra-finos partículas presentes em vesículas do retículo endoplasmático. Através de DAS-ELISA, identificou-se o Tomato chlorotic spot virus (TCSV). A partir de RNA total extraído de folhas infetadas, amplificaram-se, via RT-PCR, fragmentos correspondentes ao gene da proteína do capsídeo (cp) os quais foram seqüenciados e comparados com outros depositados no "GenBank". A homologia de nucleotídeos e aminoácidos deduzidos foi respectivamente de 99 e 95% quando comparada com seqüências de isolados de TCSV. A comparação com as outras espécies do gênero Tospovirus apresentou valores de homologia entre 72 e 84%. Estes resultados confirmam a identidade deste vírus como pertencente à espécie TCSV, que é predominante no Estado de São Paulo e importante patógeno de outras plantas cultivadas. Além disso, variedades de jiló quando inoculadas foram susceptíveis tanto ao TCSV como às espécies Tomato spotted wilt virus (TSWV) e Groundnut ringspot virus (GRSV).

A comparison of electricity spot prices simulation using ARMA-GARCH and mean-reverting models

The aim of this work is to compare two families of mathematical models for their respective capability to capture the statistical properties of real electricity spot market time series. The first model family is ARMA-GARCH models and the second model family is mean-reverting Ornstein-Uhlenbeck models. These two models have been applied to two price series of Nordic Nord Pool spot market for electricity namely to the System prices and to the DenmarkW prices. The parameters of both models were calibrated from the real time series. After carrying out simulation with optimal models from both families we conclude that neither ARMA-GARCH models, nor conventional mean-reverting Ornstein-Uhlenbeck models, even when calibrated optimally with real electricity spot market price or return series, capture the statistical characteristics of the real series. But in the case of less spiky behavior (System prices), the mean-reverting Ornstein-Uhlenbeck model could be seen to partially succeeded in this task.

Mycovellosiella robbsii sp. nov. causing leaf-spot on Mimosa caesalpiniaefolia

This is a description of the new species Mycovellosiella robsii, a cercosporoidal hyphomycete, associated with leaf spots on Mimosa caesalpiniaefolia, an important hedge plant with multiple uses in Brazil. The new species was compared to other cercosporoid species associated with plants of the genus Mimosa and other species of the genus Mycovellosiella described on legumes, and is recognized as a distinct taxon. Koch's postulates were carried out and the disease was reproduced by inoculating healthy plants with mycelial suspension or disk, confirming the pathogenicity. This is the first report of a fungal disease on this host.

Tomato chlorotic spot virus in hydroponically-grown lettuce in São Paulo State, Brazil

In the regions of Campinas and Sumaré, São Paulo, Brazil, hidroponically grown crops of Lettuce (Lactuca sativa) cv. Verônica, which showed virus-like symptoms were examined by electron microscope, biological, serological and molecular tests. Pleomorphic, enveloped particles (80-100 nm in diameter) were always detected in these samples. Experimentally inoculated host plants, including lettuce, reacted with tospoviruses-induced symptoms. Some differences were observed in Gomphrena globosa, which reacted by showing local lesions and systemic mosaic. Two isolates of Tomato chlorotic spot virus (TCSV) were identified by DAS-ELISA and by RT-PCR. The sequencing and alignment of the RT-PCR coat protein amplified fragments have indicated a high degree of homology with the TCSV sequences stored in the GenBank. This is the first report of losses due to a virus from the genus Tospovirus in commercial hydroponic lettuce crops in Brazil. Further epidemiological studies are needed for better understanding the spread of the virus in hydroponic crops, since Tomato spotted wilt virus (TSWV) is reported to spread through the nutritive solution.

Resistência de cultivares e linhagens de tomateiro a Tomato chlorotic spot virus e a Potato virus Y

Linhagens avançadas do programa de melhoramento do tomateiro (Lycopersicon esculentum) do IAC foram avaliadas em condições de campo em Campinas (SP) para resistência a tospovírus e a potyvírus, nos anos agrícolas 2002/2003 e 2003/2004, respectivamente. No primeiro ano, a única espécie de tospovírus que ocorreu na área experimental foi Tomato chlorotic spot virus (TCSV). As sete linhagens do grupo IAC exibiram baixa porcentagem de plantas sintomáticas em duas avaliações, com médias abaixo de 28%; as cultivares testadas mostraram-se altamente suscetíveis, com médias acima de 85%, à exceção de 'Franco', que apresentou cerca de 55% de infecção. No segundo experimento, conduzido em 2003/2004, dez linhagens do grupo IAC foram comparadas com cinco cultivares de polinização aberta e híbridos F1, além do acesso LA-444-1 de L. peruvianum. Nesse experimento, por meio de testes biológicos e sorológicos, verificou-se ocorrência generalizada de Potato virus Y (PVY). Foi determinado o percentual de plantas com sintomas e avaliada a intensidade dos sintomas mediante uso de escala de notas. Com base nos dois critérios, verificou-se que LA-444-1 apresenta alta resistência a PVY, que 'Tyrade' exibe comportamento intermediário, enquanto todos os demais genótipos demonstram alta suscetibilidade ao vírus. O comportamento dos genótipos avaliados neste trabalho mostra a necessidade de se considerar, nos programas de melhoramento do tomateiro, a introgressão de fatores de resistência não só a vírus de importância atual nas regiões produtoras, como geminivírus, mas também a outros vírus potencialmente nocivos à cultura, como tospovírus e potyvírus.

Effect of temperature, leaf wetness, and rainfall on the production of Guignardia citricarpa ascospores and on back spot severity on sweet orange

The black spot of citrus (Citrus sp.) is caused by Guignardia citricarpa with ascospore production depending on temperature, leaf wetness, and rainfall. The number of ascospores produced was monitored using a spore trap and climatic factors were recorded using an automated meteorological station of 'Natal' and 'Valencia' sweet orange (Citrus sinensis) orchards in Mogi Guaçu in the state of São Paulo, Brazil, from November 2000 to March 2001. The fruits were bagged to prevent infection and the bags removed from different sets of fruit for one week during each of the 18 weeks of the season in both orchards. Ascospores were produced during the entire experimental period, from spring through summer, primarily after rain events. In both orchards, ascospore production reached a peak in January and February. Ascospore production was related to leaf wetness only in the Natal orange orchard but was not related to total rainfall or temperature in either orchard. Disease was most severe on fruit exposed the 7th, 8th, and 13th weeks after beginning the experiment in both cultivars as well as after the 16th week for 'Natal'. There was a strong relationship between disease severity and total rainfall for both orchards and a weak correlation between temperature and severity in the 'Natal' block only. There was no relationship between severity and leaf wetness or ascospore numbers.