Les travaux présentés dans ce mémoire s'inscrivent dans le contexte du Groupe de Recherche en Appareillage Médical de Sherbrooke (le GRAMS), qui cherche à développer des capteurs de photons plus sensibles et plus performants, destinés à être utilisés pour détecter des photons provenant de cristaux scintillateurs notamment utilisés en tomographie d'émission par positrons. L'objectif principal du travail accompli est de faciliter la caractérisation de diodes monophotoniques à avalanche (single-photon avalanche diodes, SPAD) développées sur une technologie CMOS. Cette caractérisation couvre ce qui a trait à l'efficacité de photodétection, la résolution temporelle, les fausses détections, le redéclenchement intempestif, et la diaphonie. Un objectif optionnel est la mise au point d'un circuit réalisant la lecture d'une matrice de SPAD co-intégrée à l'aide d'un procédé d'empilement de circuits intégrés en 3D (3DIC). Ce mémoire de maîtrise présente les circuits électroniques intégrés (sur procédé CMOS 0,8μm haut voltage) et imprimés faisant partie du système électronique mis sur pied pour répondre aux objectifs du projet. Tel qu'il est démontré vers la fin du mémoire, le système a été utilisé pour caractériser des SPAD. Il a permis d'atteindre des performances dignes de l'état de l'art en circuits de contrôle de SPAD. Des améliorations au système sont proposées et seront implémentées sur des versions ultérieures.

There is a pressing need to develop alternative communications links due to a number of physical phenomena, limiting the bandwidth and energy efficiency of wire-based systems or economic factors such as cost, material-supply reliability and environmental costs. Networks have moved to optical connections to reduce costs, energy use and to supply high data rates. A primary concern is that current optical-detection devices require high optical power to achieve fast data rates with high signal quality. The energy required therefore, quickly becomes a problem. In this thesis, advances in single-photon avalanche diodes (SPADs) are utilised to reduce the amount of light needed and to reduce the overall energy budget. Current high performance receivers often use exotic materials, many of which have severe environmental impact and have cost, supply and political restrictions. These present a problem when it comes to integration; hence silicon technology is used, allowing small, mass-producible, low power receivers. A reconfigurable SPAD-based integrating receiver in standard 130nm imaging CMOS is presented for links with a readout bandwidth of 100MHz. A maximum count rate of 58G photon/s is observed, with a dynamic range of ≈ 79dB, a sensitivity of ≈ −31.7dBm at 100MHz and a BER of ≈ 1x10−9. We investigate the properties of the receiver for optical communications in the visible spectrum, using its added functionality and reconfigurability to experimentally explore non-ideal influences. The all-digital 32x32 SPAD array, achieves a minimum dead time of 5.9ns, and a median dark count rate (DCR) of 2.5kHz per SPAD. High noise devices can be weighted or removed to optimise the SNR. The power requirements, transient response and received data are explored and limiting factors similar to those of photodiode receivers are observed. The thesis concludes that data can be captured well with such a device but more electrical energy is needed at the receiver due to its fundamental operation. Overall, optical power can be reduced, allowing significant savings in either transmitter power or the transmission length, along with the advantages of an integrated digital chip.

Time resolved imaging is concerned with the measurement of photon arrival time. It has a wealth of emerging applications including biomedical uses such as fluorescence lifetime microscopy and positron emission tomography, as well as laser ranging and imaging in three dimensions. The impact of time resolved imaging on human life is significant: it can be used to identify cancerous cells in-vivo, how well new drugs may perform, or to guide a robot around a factory or hospital. Two essential building blocks of a time resolved imaging system are a photon detector capable of sensing single photons, and fast time resolvers that can measure the time of flight of light to picosecond resolution. In order to address these emerging applications, miniaturised, single-chip, integrated arrays of photon detectors and time resolvers must be developed with state of the art performance and low cost. The goal of this research is therefore the design, layout and verification of arrays of low noise Single Photon Avalanche Diodes (SPADs) together with high resolution Time-Digital Converters (TDCs) using an advanced silicon fabrication process. The research reported in this Thesis was carried out as part of the E.U. funded Megaframe FP6 Project. A 32x32 pixel, one million frames per second, time correlated imaging device has been designed, simulated and fabricated using a 130nm CMOS Imaging process from ST Microelectronics. The imager array has been implemented together with required support cells in order to transmit data off chip at high speed as well as providing a means of device control, test and calibration. The fabricated imaging device successfully demonstrates the research objectives. The Thesis presents details of design, simulation and characterisation results of the elements of the Megaframe device which were the author’s own work. Highlights of the results include the smallest and lowest noise SPAD devices yet published for this class of fabrication process and an imaging array capable of recording single photon arrivals every microsecond, with a minimum time resolution of fifty picoseconds and single bit linearity.

FLIM is branch of microscopy mainly used in biology which is quickly improving thanks to a rapid enhancement of instrumentation and techniques enabled by new sensors. In FLIM, the most precise method of measuring fluorescent decays is called TCSPC. High voltage PMT detection devices together with costly and bulky optical setups which scan the sample are usually required in TCSPC instrumentation. SPADs have enabled a big improvement in TCSPC measurement setup, providing a CMOS compatible device which can be designed in wide arrays format. However, sensors providing in-pixel TCSPC do not scale in size and in large array like the time-gated SPAD pixel sensors do. Time-gated pixels offer a less precise lifetime estimation, discarding any photon information outside a given time window, but this loss in photon-efficiency is offset by gains in pixel size. This work is aimed at the development of a wide field TCSPC sensor with a pixel size and fill factor able to reduce the cost of such devices and to obtain a high resolution time-resolved fluorescence image in the shortest time possible. The study focuses on SPAD and pixel design required to maximise the fill factor in sub 10 μm pixel pitch. Multiple pixel designs are proposed in order to reduce pixel area and so enable affordable wide array TCSPC systems. The first proposed pixel performs the CMM lifetime estimation in order to reduce the frame rate needed to stream the data out of the SPAD array. This pixel is designed in a 10 μm pitch and attains with the most aggressive design a fill factor of 10:17 %. A second design proposes an analogue TCSPC which consists in a S/H TAC circuitry. This simpler pixel can achieve a higher fill factor of 19:63% as well as smaller pitch of 8 μm thanks to the adoption of SPAD n-well and electronics area sharing. This last design is implemented in a 320 x 256 SPAD array in which is included part of a novel ADC aimed at reduction of the processing time required to build a TCSPC histogram. A more conventional analogue readout is used to evaluate the pixel performance as well as a more fine TCSPC histogram. The device was used to measure the fluorescence lifetime of green micro-spheres while the 2b flash ADC is used to demonstrate rapid resolution and separation of two different fluorescence decays.

Submitted by Fabiana Morbi Fernandes null (fabianamorbi90@gmail.com) on 2017-08-18T11:48:51Z No. of bitstreams: 1 Tese Fabiana-versao Final 2.pdf: 1313330 bytes, checksum: ead7f093a270db6026255c4abdbfe37c (MD5) / Approved for entry into archive by Luiz Galeffi (luizgaleffi@gmail.com) on 2017-08-23T18:18:29Z (GMT) No. of bitstreams: 1 fernandes_fm_me_bot.pdf: 1313330 bytes, checksum: ead7f093a270db6026255c4abdbfe37c (MD5) / Made available in DSpace on 2017-08-23T18:18:30Z (GMT). No. of bitstreams: 1 fernandes_fm_me_bot.pdf: 1313330 bytes, checksum: ead7f093a270db6026255c4abdbfe37c (MD5) Previous issue date: 2017-07-21 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O nitrogênio (N) é um dos nutrientes extraídos em maior quantidade pela batateira e tem grande impacto no desempenho dessa cultura. A resposta da cultura da batata à adubação nitrogenada é dependente de diversos fatores, o que torna essencial o manejo correto do N, maximizando assim a absorção deste pela planta. Além disso, a aplicação de doses insuficientes ou demasiadamente elevadas de N pode reduzir a produtividade e a qualidade dos tubérculos produzidos e representar prejuízos aos produtores e riscos ao ambiente. Neste sentido, a estimativa da necessidade de N, mediante leituras do índice relativo de clorofila (IRC) nas folhas da cultura, com o clorofilômetro portátil, pode ser uma alternativa viável para tornar o manejo do N mais preciso. Contudo, ainda existe a necessidade de estabelecimento de critérios para a utilização dessa ferramenta na cultura da batata. Dessa forma, os objetivos deste trabalho foram: a) avaliar a correlação entre o IRC, obtido mediante leituras do clorofilômetro portátil, e o teor de N da folha de duas cultivares de batata (Agata e Electra), em diversos estádios de desenvolvimento e b) avaliar qual valor (90% e 95%) do índice de suficiência de N (ISN), calculado com base nas medidas IRC nas folhas em relação ao tratamento referência, é o mais adequado para indicar o momento da adubação nitrogenada em duas cultivares de batata e, consequentemente, proporcionar maior produtividade, qualidade de tubérculos e eficiência de utilização do N aplicado. Durante a safra “de inverno” de 2016, foram realizados quatro experimentos, sendo dois na Fazenda Experimental Lageado (Local 1) e dois em área particular de produção de batata (Local 2), ambas no município de Botucatu-SP. Em cada local foi conduzido um experimento com cada cultivar. Em todos os experimentos, o delineamento experimental utilizado foi em blocos casualizados, com quatro repetições. Os tratamentos foram constituídos por seis manejos do N: M1: 80 kg ha-1 de N no sulco de plantio + 80 kg ha-1 de N aos 10 dias após a emergência (DAE) + 80 kg ha-1 de N aos 30 DAE + 80 kg ha-1 de N aos 45 DAE (referência); M2: 80 kg ha-1 de N no sulco de plantio + 80 kg ha-1 de N aos 10 DAE (recomendado); M3: 160 kg ha-1 de N no sulco de plantio (baseado na adubação comumente utilizada pelos produtores, simulando aplicação de 4.000 kg ha-1 da fórmula N-P2O5-K2O 04-14-08); M4: 60 kg ha-1 de N no sulco de plantio + 40 kg ha-1 de N quando as leituras do clorofilômetro indicarem ISN < 90%; M5: 60 kg ha-1 de N no sulco de plantio + 40 kg ha-1 de N quando as leituras do clorofilômetro indicarem ISN < 95% e M6: testemunha (sem aplicação de N). O IRC, obtido mediante as leituras do clorofilômetro, correlacionou-se de forma positiva com o teor de N na folha das cultivares de batata Agata e Electra a partir dos 24 DAE. A aplicação de maiores doses de N, especialmente de forma parcelada, proporcionou maiores quantidades de N acumuladas e maior produtividade de tubérculos pela batateira, especialmente no Local 2 para ambas as cultivares, sendo o clorofilômetro mais preciso para indicar a ausência de deficiência de N, do que o momento e a necessidade de realização da adubação nitrogenada. O uso do ISN<90%, baseado na medida do clorofilômetro em uma área referência, permitiu definir quando se deve aplicar o N em cobertura na cultura da batata nas cultivares Agata e Electra, melhorando a eficiência de utilização do N, sendo este o capaz de proporcionar economia de N aplicado sem ocasionar redução na produtividade de tubérculos, quando comparadas com o tratamento referência (sem deficiência de N) e com a dose de N recomendada para a cultura. / Nitrogen (N) is one of the nutrients extracted in greater quantity by potato and has a great impact on the performance of this crop. The response of the potato crop to the nitrogen fertilization is dependent on several factors, which makes essential the correct management of N, thus maximizing its absorption by the plant. In addition, the application of insufficient or excessively high doses of N may reduce the productivity and quality of the tubers produced and represent damages to the producers and risks to the environment Therefore, the estimation of the need for N, through readings of the relative index of chlorophyll (IRC) in the leaves of the culture, with the portable chlorophyllometer, can be a viable alternative to make N management more accurate. However, there is still a need to establish criteria for the use of this tool in the potato crop. Thus, the objectives of this study were: a) evaluate the correlation between the IRC, obtained by readings of chlorophyll, and the N concentration in leaves of two potato cultivars (Agata e Electra) in various stages of development, and b) evaluate what value (90% or 95%) of N sufficiency index (NSI), calculated based on RCI measures in the leaves regarding the treatment reference, is best suited to indicate the time of N fertilizer in two cultivars potato and hence to higher tuber yields and quality, and N-use efficiency. In the year 2016, during winter season four experiments were carried out in the field conditions, two at Lageado Experimentl Farm (Site 1) and two in a private potato farm (Site 2), both in the municipality of Botucatu. In each site were carried out one experiment with each cultivar. The design used in each experiment will be randomized blocks, with six N managements (M1: 80 kg ha-1 N in the planting furrow + 80 kg ha-1 at 10 days after emergence (DAE) + 80 kg ha-1 at 30 DAE + 80 kg ha-1 at 45 DAE (reference); M2: 80 kg ha-1 N in the furrow + 80 kg ha-1 at 10 DAE (recommended) M3: 160 kg ha-1 N in the furrow (representing the application of 4000 kg ha-1 N-P2O5-K2O formula 04-14-08); M4: 60 kg ha-1 N in the furrow + 40 kg ha-1 when the chlorophyll readings indicate NSI < 90%; M5: 60 kg ha-1 N in the furrow + 40 kg ha-1 when the chlorophyll readings indicate NSI < 95% and M6: control - without N application) and four replications. The RCI, obtained through the chlorophyllometer readings, was positively correlated with the N content in the leaves of the Agata and Electra potato cultivars from 24 DAE. The application of higher doses of N, especially in a piecemeal manner, provided higher amounts of N accumulated and higher yield of tubers of potato, especially in Site 2 for both cultivars, being the chlorophyllometer more accurate to indicate the absence of N deficiency, than the time and need for nitrogen fertilization. The use of the NSI <90%, based on the measurement of the chlorophyllometer in a reference area, allowed to define when N to cover the potato crop in the Agata and Electra cultivars should be applied, improving the efficiency of N utilization, of providing N economy applied without causing a reduction in tuber yield when compared to the reference treatment (without N deficiency) and to the N dose recommended for cultivation.

Rose-scented geranium (Pelargonium capitatum x P. radens) belongs to the family Geraniaceae and it is a multi-harvest, high value, commercially important essential oil yielding aromatic plant. The essential oil extracted from the herbage of the plant is widely used in the fragrance and cosmetics industry and scenting of soaps. The essential oil is extracted by steam distillation. South Africa is now producing significant quantities of geranium oil. However, previous experience by this research group showed that seedlings often take long to establish, resulting in high death rates and sometimes poor growth after establishment. Stunted growth and yellowing of leaves was also observed in some cases. Poor vegetative growth causes low herbage yield and, consequently, low total essential oil production per hectare. Poor growth is believed to be due to a combination of factors, including nutrient deficiencies and acidic soil conditions and has not been studied under South Africa condition. Production of the crop is also reported to respond differently to rate and source of nutrition in various agro-ecological regions of the world. Therefore, field trials were conducted at the Hatfield Experimental Farm, University of Pretoria, in order to investigate how the plant responds to agronomic practices, such as source and amount of nitrogen, time of N fertilizer application, season of N fertilization and liming. Response of rose-scented geranium to source and amount of N showed that, at the first harvest (summer/autumn), there was no significant effect of conventional N on fresh herbage and oil yield, probably due to leaching of N by rainfall. However, organic N at 100 kg•ha-1 increased fresh herbage and oil yields by 58% and 48% over the control, respectively. In the second harvest (spring/summer), fresh herbage yield increased by 46% (conventional N) and 60% (organic N) at 100 kg•ha-1 compared to the control. Compared to the control, 100 kg•ha-1 conventional and organic N also increased essential oil yields by 94% and 129%, respectively. For both N sources nitrogen use efficiency (NUE) and LAI decreased with an increase in N level, and organic N gave highest essential oil production efficiency and LAI. Essential oil content (% fresh mass basis) also varied between the harvests, being greater in the second harvest (September to December 2005; spring/summer) than the first harvest (February to May 2005; summer/autumn). This was due to environmental variations that occurred between the harvesting periods. N level and source were found to have no noticeable effect on essential oil composition. This study revealed that rose-scented geranium produced higher fresh herbage and essential oil yield when organic fertilizer was used as a source of N. Nitrogen management in terms of rate and time of application is important in rose-scented geranium production. Delaying nitrogen topdressing (conventional N in the form of LAN; N 28%) after harvest to between the 7th and 9th week after cut back, was found to have a significant positive effect on biomass and essential oil production. Essential oil content of the plant did not show any response to a delay in nitrogen topdressing. A delay in nitrogen topdressing, in the first re-growth resulted into a lower citronellol to geraniol (C:G) ratio, which favour essential oil quality of the crop. Generally, the characteristics of the essential oil were within the internationally acceptable range for rose-scented geranium essential oil. In addition, production of rose-scented geranium during cooler periods is not advisable due to limited biomass production which might encourage leaching of nitrogen. The net benefits from N application is dependent on the growing period and in the present study spring and summer were more beneficial than winter and autumn. Organic N at 100 kg•ha-1•year-1 increased herbage and essential oil yield of the crop in spring and summer but further increases in organic or conventional N levels had no significant effect. N application either in winter or autumn did not improve production of the crop. Application of more N than what is required for optimum growth of the plant had no positive effect on essential oil production. Application of organic N also resulted in higher N use efficiency than conventional N. The essential oil contents (% fresh mass basis) achieved in the present study generally fell within the range of 0.04 to 0.2%. Citronellol and geraniol concentration (%), were at peak in spring season followed by summer and autumn and lowest in winter. The ratio between these two components (C:G ratio) is also used as an indicator of rose-scented geranium essential oil quality and most desirable (low C:G ratio) essential oil was attained in spring, summer and autumn harvesting seasons and least desirable oil was attained in winter. The relationship between SPAD-502 chlorophyll meter readings (SPAD units) and leaf N content (% dry weight basis) was a quadratic function. SPAD-502 chlorophyll meter readings (SPAD units) matched well with that of leaf N concentration data of rose-scented geranium. Regardless of the factors that affect the readings, this instrument can be used as an indicator of leaf N status of rose-scented geranium. Soil pH above 5.5 and soil base saturation above 55% increased fresh herbage and essential oil yield (per ha), which corresponded in this case with 2 to 6 t•ha-1 of lime application. Oil content (%) was not significantly affected by application of lime. Therefore, optimum growth of rose-scented geranium can be achieved by application of lime when plants are grown on acidic soils, but without any effect on oil content and essential oil composition. It can be concluded that N rate, source and season of production and soil pH should be considered to ensure optimal rose-scented geranium production. / Thesis (PhD)--University of Pretoria, 2012. / Plant Production and Soil Science / PhD / Unrestricted

Abstract Endive (Cichorium endivia L.) is a leafy vegetable mainly used in ready-to use salads. Nitrate accumulation, is a serious problem in this type of vegetables, because when ingested, nitrate has toxic effects in the human health. Solar radiation, nitrogen fertilization and temperature are some factors that affect nitrate concentration in leaves. 'Tipburn' , bolting and 'glassines' are some physiological disorders that appears in endives and impairs their marketable quality. 'Tipburn' is characterized by necrosis on the outside of leaves, and 'glassines' is associated with oil spots on leaves. In this thesis have been studied the influence of different environmental factors (temperature, relative humidity), nitrogen fertilization and cultivar in the nitrate accumulation, the incidence of physiological disorders and marketable production. Also, we studied the correlations between different parameters in order to know, with some of this, if the chlorophyll meters SPAD-502 and the colorimeter could be useful for estimating indirectly nitrate and nitrogen content in endives. Different experiments were realized using broad-leaved endive cultivars ('Cuartana', 'Natacha', 'Santel') that were cultivated in open-field and in greenhouse with different humidity levels in different seasons. Regarding nitrogen fertilization, the experiments were separated in two groups. In the first group the NH4+/NO3- ratio was modified, increasing the percentage of nitrogen applied in ammonium form (0, 5, 10, 20%). In the second, three nutrient solutions were applied. In these solutions, a part of the nitrates were replaced by chloride (0, 20, 40% of total nitrogen). Periodically, the chlorophyll content (extracted by acetone and measured with a chlorophyll meter SPAD-502), color parameters, nitrate concentration (reflectometry), total nitrogen (Kjeldahl) and phosphorus were determined. Also nitrate reductase activity was measured. At the end of each cycle the production and the incidence of physiological disorders were determined. Results showed that cv. 'Cuartana' was more resistant to 'tipburn'. The hotest cycles and greenhouse favored the incidence of 'tipburn', but increasing the relative humidity we achieved reduced it. Cv. 'Santel' presented the highest susceptibility to bolting in conditions of short photoperiod. 'Glassines' was favored by low relative humidity, and cv. 'Cuartana' was the most susceptible. Nitrate accumulation was higher in spring-summer cycles and in greenhouse. Applying a 10-20% of ammonium with the nutrient solution and replacing part of the nitrates by chloride, nitrate concentration in endive leaves was reduced, maintaining the quality of the plants. In the other hand, in our conditions, the chlorophyll meter SPAD-502 and the colorimeter wouldn't be useful for estimate indirectly the nitrate and nitrogen content in plants. / Resumen La escarola (Cichorium endivia L.) es una hortaliza aprovechable por sus hojas tanto de forma directa como en presentaciones de 'IV gama'. La acumulación de nitratos en este tipo de hortalizas es un factor a tener en cuenta en su cultivo, puesto que su ingesta tiene efectos perjudiciales en la salud humana y que se encuentra influenciada por factores como la fertilización nitrogenada, la radiación y la temperatura. El 'tipburn', la subida a flor prematura y la 'mancha translúcida' son tres de las fisiopatías que afectan a la calidad comercial de la escarola. El 'tipburn' se manifiesta como una desecación marginal de las hojas, mientras que la 'mancha translúcida' se presenta como manchas aceitosas en los limbos. En esta tesis doctoral se ha estudiado la influencia de diferentes factores ambientales (temperatura y humedad), de la fertilización nitrogenada y del cultivar empleado en la acumulación de nitratos, incidencia de fisiopatías y producción comercial en plantas de escarola. Por otro lado, se han establecido correlaciones entre los diferentes parámetros fisiológicos determinados con el fin de conocer, a través de algunas de ellas, si los medidores portátiles SPAD-502 y el colorímetro podrían emplearse para determinar de forma indirecta el contenido en nitratos y nitrógeno. Para ello se realizaron diferentes experimentos utilizando en todos ellos cultivares de escarola de hoja lisa ('Cuartana', 'Natacha', 'Santel') que fueron cultivados tanto al aire libre como bajo invernadero y bajo distintos niveles de humedad en diferentes ciclos de cultivo. Respecto de la fertilización, los experimentos se dividieron en dos bloques. En el primero de ellos se realizaron varios ensayos en los cuales se modificó la ratio NH4+/NO3-, variando el porcentaje de nitrógeno aportado en forma de amonio (0, 5, 10 y 20%). En el segundo bloque, se aportaron tres soluciones nutritivas, en las que parte de los nitratos se reemplazaron por iones cloruro representando un 0, 20 y 40% respecto del nitrógeno total. Periódicamente se determinó el contenido en clorofilas (extracción mediante acetona y medidor SPAD-502), color, concentración de nitratos (reflectometría), porcentaje de nitrógeno total (Kjeldahl) y fósforo, así como la actividad de la enzima nitrato-reductasa. Al final del ciclo se determinó la producción y la incidencia de fisiopatias. Los resultados mostraron que el cultivar 'Cuartana' resultó más resistente a la aparición de 'tipburn', cuya incidencia fue más severa en ciclos estivales y bajo invernadero, pero se consiguió disminuirla con incrementos de la humedad relativa. El cultivar 'Santel' presentó una mayor susceptibilidad a la subida a flor prematura, que afectó en mayor medida en condiciones de fotoperiodo corto, mientras que la 'mancha traslucida' se vio favorecida por las condiciones de humedad relativa más baja, siendo más susceptible el cultivar 'Cuartana'. La acumulación de nitratos fue superior en ciclos primaverales-estivales y bajo invernadero. Con el aporte de un 10-20% de amonio en la solución nutritiva y el reemplazo de parte de los nitratos por iones cloruro se redujo el contenido de nitratos de las hojas de escarola sin verse disminuida su calidad comercial. Por otro lado, en nuestras condiciones el medidor SPAD-502 así como el colorímetro no serían herramientas útiles para estimar de manera indirecta el contenido en nitratos y nitrógeno de la planta. / Resum L'escarola (Cichorium endivia L.) és una hortalissa aprofitable per les seues fulles, tant de forma directa, com en presentacions de 'IV gama'. L'acumulació de nitrats en aquest tipus d'hortalisses, és un factor a tenir en compte en el seu cultiu, ja que la seua ingesta té efectes perjudicials en la salut humana, i es trova influenciada per factors com la fertilització nitrogenada, la radiació i la temperatura. El 'tipburn', la floració prematura i la 'taca translúcida', són tres de les fisiopaties que afecten a la qualitat comercial de l'escarola. El 'tipburn' es manifesta com una dessecació marginal de les fulles, mentre que la 'taca translúcida' es presenta en forma de taques olioses als limbes. A aquesta tesi doctoral s'ha estudiat la influència de diferents factors ambiental, como la temperatura i la humitat relativa, així com de la fertilització nitrogenada i del cultivar emprat en l'acumulació de nitrats, incidència de fisiopaties i producció comercial en plantes d'escarola. Per altra banda, s'han establert correlacions entre els diferents paràmetres fisiològics determinats, amb l'objectiu de conèixer, mitjançant algunes d'elles, si els mesuradors portàtils de clorofil·la SPAD-502 i el colorímetre es podrien utilitzar per a determinar de forma indirecta el contingut en nitrats i nitrogen total. Per assolir aquestos objectius, es varen realitzar diferents experiments emprant en tots ells cultivars d'escarola de fulla llisa ('Cuartana', 'Natacha', 'Santel') que foren cultivats tant a l'aire lliure con a l'hivernacle, amb distints nivells d'humitat en diferents cicles de cultiu. Quan a la fertilització nitrogenada, els experiments es varen dividir en dos grups. En el primer d'ells es varen dur a terme diferents assaigs en els quals, es va modificar la ratio NH4+/NO3- variant el percentatge de nitrogen aplicat en forma d'amoni (0, 5, 10 y 20%). En el segon bloc, s'aportaren tres solucions nutritives, en les quals part dels nitrats van ser reemplaçats per ions clorur, representant un 0,20 y 40% respecte del nitrogen total. Periòdicament es va determinar el contingut en clorofil·les (mitjançant extracció amb acetona i amb el mesurador SPAD-502), el color, la concentració de nitrats (reflectometria), percentatge de nitrogen (Kjeldahl) i fòsfor, així com l'activitat de l'enzima nitrat-reductasa. A la fi de cada cicle de cultiu es va determinar la producció i la incidència de fisiopaties. Els resultats mostraren que el cultivar 'Cuartana' va ser més resistent a l'aparició de 'tipburn'. Fisiopatia que va aparèixer amb més intensitat en cicles estivals i en hivernacle, però que es va reduir amb increments de la humitat relativa. El cultivar 'Santel' presentà una major susceptibilitat a la floració prematura, que va afectar en major mesura en condiciones de fotoperíode curt. La 'taca translúcida' es va veure afavorida per les condicions d'humitat relativa més baixa, sent més susceptible el cultivar 'Cuartana'. L'acumulació de nitrats va ser superior en els cicles de primavera i estiu i en el cultiu a l'hivernacle. Amb l'aplicació d'un 10-20% d'amoni en la solució nutritiva i el reemplaçament de part dels nitrats per ions clorur, es va reduir el contingut de nitrats en les fulles d'escarola, sense veure un descens en la qualitat comercial. D' altra banda, en les nostres condiciones, el mesurador SPAD-502, així com el colorímetre, no serien unes eines útils per estimar de manera indirecta el contingut en nitrats i nitrogen de la planta. / Gromaz Roig, A. (2017). Escarola (Cichorium endivia L.): sistemas de manejo y ciclos como factores de la incidencia de fisiopatías y la acumulación de nitratos [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/78295

Plantas cultivadas em ambientes com média e alta disponibilidade de cádmio apresentam menor produção de folhas e perfilhos devido à maior quantidade de cádmio absorvido, resultando em menor produção de biomassa. O fornecimento de enxofre às plantas pode minimizar os efeitos negativos causados pelo cádmio, uma vez que esse nutriente participa de compostos que atuam no sistema antioxidante, conferindo maior tolerância aos metais pesados. Objetivou-se avaliar as modificações ocorridas: i) nos aspectos morfológicos, morfogênicos e produtivos; ii) no estado nutricional e iii) na resposta do sistema antioxidante do capim-tanzânia (Panicum maximum) cultivado com combinações de doses de enxofre (0,1; 1,0; 1,9; 2,8 e 3,7 mmol L-1) e de cádmio (0,0; 0,5; 1,0; 1,5 e 2,0 mmol L-1) em solução nutritiva. O experimento foi desenvolvido em casa de vegetação, utilizando o esquema fatorial 5² fracionado, em blocos ao acaso, com seis repetições. O primeiro corte do capimtanzânia foi realizado 40 dias após o transplantio das mudas para os vasos e o segundo corte 18 dias após o primeiro. Para a taxa de mortalidade de perfilhos, valor SPAD, concentração de cádmio e a atividade da APX (segundo corte) do capim-tanzânia foi significativa a interação doses de enxofre x doses de cádmio, de forma que o suprimento de enxofre atenuou a taxa de mortalidade, diminuiu a quantidade de cádmio absorvida, aumentou o valor SPAD e a atividade da APX. O teor de água, número de perfilhos, taxa de aparecimento de perfilhos, número de folhas, área foliar, taxa de aparecimento foliar, razão de área foliar, produção de massa seca da parte aérea e das raízes, valor SPAD, concentração de potássio, teor de GSH e a atividade da CAT foram menores com o fornecimento de cádmio na solução nutritiva. De forma contrária, as concentrações de nitrogênio, fósforo, zinco e cádmio, o fator de transporte do cádmio, os teores de GSSG e glutationa, e a atividade da APX aumentaram. O fornecimento de enxofre aumentou a concentração de enxofre na parte aérea do capimtanzânia. Os efeitos causados pelo cádmio são mais acentuados durante a rebrotação, principalmente, em condições de alta disponibilidade do metal pesado. As enzimas CAT e GR apresentaram mais isoformas na parte aérea do que nas raízes do capim-tanzânia, sugerindo que esse tecido é mais eficaz no combate aos danos oxidativos causados pelas altas concentrações de cádmio. O suprimento de enxofre ao capim-tanzânia cultivado em ambientes com alta disponibilidade de cádmio é indicado para aumentar a tolerância da gramínea ao metal pesado. / Plants grown at medium and high cadmium availability has produced low number of leaves and tillers due to the great amount of cadmium absorbed, resulting in lower biomass production. Sulfur supply to plants may mitigate negative effects caused by cadmium, since compounds containing sulfur participate of the antioxidative plant system, resulting in greater tolerance to heavy metals. This study evaluated changes in: i) morphological, morphogenesis and productive aspects, ii) nutritional status, and iii) response of the antioxidative system of Tanzania guineagrass (Panicum maximum) grown under combined sulfur rates (0.1; 1.0, 1.9, 2.8 and 3.7 mmol L-1) and cadmium rates (0.0, 0.5, 1.0, 1.5 and 2.0 mmol L-1) in nutrient solution. The experiment was conducted in greenhouse, by using a fractionated factorial 5², in randomized block design, with six replications. The first harvest of Tanzania guineagrass was performed at 40 days after seedlings transplanting to the pots, and the second at 18 days after the first harvest. Mortality rate of tillers, SPAD value, cadmium concentration and APX activity (second harvest) of Tanzania guineagrass had statistically significant sulfur rates x cadmium rates interaction, since that sulfur supply mitigate the mortality rate, decreased the amount of cadmium absorbed and increased SPAD value and the APX activity. The water content, number of tillers, tiller emergence rate, number of leaf, leaf area, leaf emergence rate, ratio of leaf area, dry mass of shoots and roots, SPAD value, potassium concentration, GSH content and CAT activity decreased with the cadmium supply in the nutrient solution. Contrary to that, nitrogen, phosphorus, zinc and cadmium concentrations were increased as well as the transporting factor of cadmium, GSSG and glutathione contents and APX activity. Sulfur supply increased sulfur concentration in shoots of Tanzania guineagrass and the effects of cadmium were more pronounced during the plant regrowth period, especially under high availability of the heavy metal. Isoforms of CAT and GR showed higher abundance in shoots compared to roots, suggesting higher effectiveness of such tissue in avoiding oxidative damage caused by high concentrations of cadmium. Thus, sulfur supply to the Tanzania guineagrass grown in environment with high cadmium availability is indicated to increase tolerance to this heavy metal.

Plantas cultivadas em ambientes com média e alta disponibilidade de cádmio apresentam menor produção de folhas e perfilhos devido à maior quantidade de cádmio absorvido, resultando em menor produção de biomassa. O fornecimento de enxofre às plantas pode minimizar os efeitos negativos causados pelo cádmio, uma vez que esse nutriente participa de compostos que atuam no sistema antioxidante, conferindo maior tolerância aos metais pesados. Objetivou-se avaliar as modificações ocorridas: i) nos aspectos morfológicos, morfogênicos e produtivos; ii) no estado nutricional e iii) na resposta do sistema antioxidante do capim-tanzânia (Panicum maximum) cultivado com combinações de doses de enxofre (0,1; 1,0; 1,9; 2,8 e 3,7 mmol L-1) e de cádmio (0,0; 0,5; 1,0; 1,5 e 2,0 mmol L-1) em solução nutritiva. O experimento foi desenvolvido em casa de vegetação, utilizando o esquema fatorial 5² fracionado, em blocos ao acaso, com seis repetições. O primeiro corte do capimtanzânia foi realizado 40 dias após o transplantio das mudas para os vasos e o segundo corte 18 dias após o primeiro. Para a taxa de mortalidade de perfilhos, valor SPAD, concentração de cádmio e a atividade da APX (segundo corte) do capim-tanzânia foi significativa a interação doses de enxofre x doses de cádmio, de forma que o suprimento de enxofre atenuou a taxa de mortalidade, diminuiu a quantidade de cádmio absorvida, aumentou o valor SPAD e a atividade da APX. O teor de água, número de perfilhos, taxa de aparecimento de perfilhos, número de folhas, área foliar, taxa de aparecimento foliar, razão de área foliar, produção de massa seca da parte aérea e das raízes, valor SPAD, concentração de potássio, teor de GSH e a atividade da CAT foram menores com o fornecimento de cádmio na solução nutritiva. De forma contrária, as concentrações de nitrogênio, fósforo, zinco e cádmio, o fator de transporte do cádmio, os teores de GSSG e glutationa, e a atividade da APX aumentaram. O fornecimento de enxofre aumentou a concentração de enxofre na parte aérea do capimtanzânia. Os efeitos causados pelo cádmio são mais acentuados durante a rebrotação, principalmente, em condições de alta disponibilidade do metal pesado. As enzimas CAT e GR apresentaram mais isoformas na parte aérea do que nas raízes do capim-tanzânia, sugerindo que esse tecido é mais eficaz no combate aos danos oxidativos causados pelas altas concentrações de cádmio. O suprimento de enxofre ao capim-tanzânia cultivado em ambientes com alta disponibilidade de cádmio é indicado para aumentar a tolerância da gramínea ao metal pesado. / Plants grown at medium and high cadmium availability has produced low number of leaves and tillers due to the great amount of cadmium absorbed, resulting in lower biomass production. Sulfur supply to plants may mitigate negative effects caused by cadmium, since compounds containing sulfur participate of the antioxidative plant system, resulting in greater tolerance to heavy metals. This study evaluated changes in: i) morphological, morphogenesis and productive aspects, ii) nutritional status, and iii) response of the antioxidative system of Tanzania guineagrass (Panicum maximum) grown under combined sulfur rates (0.1; 1.0, 1.9, 2.8 and 3.7 mmol L-1) and cadmium rates (0.0, 0.5, 1.0, 1.5 and 2.0 mmol L-1) in nutrient solution. The experiment was conducted in greenhouse, by using a fractionated factorial 5², in randomized block design, with six replications. The first harvest of Tanzania guineagrass was performed at 40 days after seedlings transplanting to the pots, and the second at 18 days after the first harvest. Mortality rate of tillers, SPAD value, cadmium concentration and APX activity (second harvest) of Tanzania guineagrass had statistically significant sulfur rates x cadmium rates interaction, since that sulfur supply mitigate the mortality rate, decreased the amount of cadmium absorbed and increased SPAD value and the APX activity. The water content, number of tillers, tiller emergence rate, number of leaf, leaf area, leaf emergence rate, ratio of leaf area, dry mass of shoots and roots, SPAD value, potassium concentration, GSH content and CAT activity decreased with the cadmium supply in the nutrient solution. Contrary to that, nitrogen, phosphorus, zinc and cadmium concentrations were increased as well as the transporting factor of cadmium, GSSG and glutathione contents and APX activity. Sulfur supply increased sulfur concentration in shoots of Tanzania guineagrass and the effects of cadmium were more pronounced during the plant regrowth period, especially under high availability of the heavy metal. Isoforms of CAT and GR showed higher abundance in shoots compared to roots, suggesting higher effectiveness of such tissue in avoiding oxidative damage caused by high concentrations of cadmium. Thus, sulfur supply to the Tanzania guineagrass grown in environment with high cadmium availability is indicated to increase tolerance to this heavy metal.

Le noyau d'une cellule est hétérogène par sa structure et son activité et beaucoup de ses composants interagissent de façon dynamique. Lors de l'étude de processus cellulaires comme la réponse au stress thermique, des expériences classiques de spectroscopie de corrélation de fluorescence (FCS), qui sont habituellement limitées à un seul volume d'observation, n'apportent que des résultats partiels à cause des informations spatiales manquantes. Ce mémoire de thèse présente une nouvelle technique de FCS multi-confocale (mFCS) qui permet des mesures FCS simultanées à différents endroits d'une cellule. La technique est basée sur l'emploi d'un modulateur spatial de lumière pour la création de plusieurs volumes d'observations distincts et d'une caméra ``electron-multiplying'' CCD (EMCCD) pour la détection en parallèle. La résolution spatiale ainsi que la sensibilité du système mFCS sont proches de celles d'un système FCS classique et en utilisant un mode d'acquisition particulier une résolution temporelle de $14mu s$ a pu être atteinte. La technique mFCS est appliquée à l'étude de la réponse cellulaire au stress thermique en observant le facteur de transcription heat shock factor 1 (HSF1), qui est un régulateur clé de la réponse au stress thermique. Des mesures mFCS dans des cellules vivantes révèlent des changements dans la dynamique de HSF1 pendant le choc thermique. Ces changements concernent l'affinité ainsi que l'homogénéité spatiale des interactions avec l'ADN. En outre, nous avons également évalué les performances d'une caméra CMOS-SPAD et testé le dispositif en tant que capteur alternatif pour la mFCS en cellules vivantes. / The cell nucleus is heterogeneous in its structure and activity and many of its components are in dynamic interactions with each other. When investigating the cellular response to an external signal, such as heat shock, standard fluorescence correlation spectroscopy (FCS) experiments, which are limited to one observation volume, do only give partial results because of the missing spatial information. This work introduces a novel multi-confocal FCS (mFCS) technique that allows simultaneous FCS measurements in different locations within a cell. It is based on the use of a spatial light modulator (SLM) to create several distinct observation volumes at a time and an electron-multiplying charge coupled device (EMCCD) camera to perform parallel detection. The spatial resolution as well as the sensibility of the mFCS system are close to that of a classical FCS setup and using a special readout mode, a temporal resolution of $14mu s$ is reached. The mFCS technique is applied to study the cellular response to thermal stress by monitoring the transcription factor heat shock factor 1 (HSF1), which is a key regulator of the heat shock response. mFCS experiments in living cells reveal changes in the dynamics of HSF1 upon heat shock. These changes concern the affinity as well as the spatial homogeneity of its interactions with DNA. Additionally, the performance of a CMOS-SPAD camera, consisting of an array of single photon avalanche diodes, is evaluated and the device is tested as an alternative detector for mFCS in living cells.