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Introduction: Part 1. Greenhouse studies were conducted to investigate the absorption of sodium, potassium, calcium and magnesium by lemon seedlings. The plants were grown in controlled nutrient solutions and analysis of the plant material was made to determine the relationship between the four cations in the different plant organs. Results of the experiment may be summarized as follows:- The occurrence of leaf burn appeared to be associated with a low calcium content, together with a high sodium, potassium or sodium + potassium content. The development of the seedlings was shown to be markedly influenced by the level of calcium supply and to a much lesser degree by the level of potassium supplied. The concentration of sodium, potassium, calcium and magnesium in the stem and leaf varied with position of the tissues on the main axis. The level of sodium was found to influence the distribution of sodium in the leaves, and the sodium content of the leaf, stem and root tissues gave a good reflection of the level of sodium supplied. The effect of the sodium treatment on the uptake of potassium appeared to be dependent on the calcium content of the tissues. Sodium treatment was found not to affect the leaf calcium content. In the stem and root tissues the calcium content was reduced when the calcium level in the nutrient medium was low, and increased the calcium content of these two tissues at the higher level of calcium supply. The sodium supply was shown to reduce the leaf magnesium only slightly, and its affect on the magnesium content of the stem and root tissues was shown to be dependent on the level of calcium supplied. The potassium content of the leaves, stems and roots increased when the supply of potassium was raised. Increasing the potassium level in the nutrient supply was found to cause a very slight reduction in the sodium content of the leaf, but did not affect the sodium content of the stem and root. Potassium did not affect the calcium content of the leaves and increased the stem calcium when sodium was not present in the nutrient media in high concentrations. In the roots potassium only increase the calcium content at the upper level of calcium supply. The magnesium content of the leaf and stem tissues was reduced as the potassium supply was increased. In the roots the general trend was for the magnesium content to increase when the level of potassium treatment was raised. The calcium content of all the plant parts increased with a rise in the calcium supply, the stem and root tissues having a greater percentage increase than the leaves. The sodium content of the plant tissue was reduced as the level of calcium supply was raised. The leaf potassium content decreased with a higher calcium level of supply. In the stem and roots the potassium content was increased by doubling the calcium supply. The calcium treatment had no influence on the magnesium content of the foliage, but in the stem and root tissues a decrease in magnesium content resulted from an increase in the calcium supply. Part II Potato tuber tissue and carrot root tissue were used as experimental material in the investigation of sodium9 potassium9 and calcium uptake by storage tissue. It was shown that when the tissue disks are transferred from distilled water to a solution of salts there is a rapid initial uptake of cation which is neither particularly selective, nor related to metabolism, but dependent on the external concentration. On the other hand, the prolonged active accumulation of cation exhibits selectivity. Potassium absorption by potato tuber tissue was shown to be stimulated by sodium9 whereas the potassium and calcium absorption by carrot root tissue was shown to be reduced by sodium. Similarly the sodium absorption by the carrot root tissue was reduced by potassium, thus suggesting that the cations compete with one another for the same absorption mechanism.

The taxonomy of Fusarium species has been controversial. The controversy surrounding the morphology-based classification, as well as the limitations and advantages of the various tools used to circumvent this problem are discussed as an introduction to the research presented in this thesis. Emphasis in this case was on the F. oxysporum species complex. The body of the research focuses on Fusarium species in Ethiopian agriculture about which little has been published. Results from this study suggest that F. oxysporum and F. solani are the two species most commonly isolated from agricultural soils and plant tissues from Ethiopia. The Ethiopian F. solani isolates were further characterized using AFLPs which showed that these isolates group into two separate clusters. Additional characterization using EF-1á sequence analyses showed that all these isolates belong to one of the three clades previously designated for the F. solani-Nectria haematococca species complex. Nine SSR markers were developed for studying the F. oxysporum complex. These markers are highly polymorphic and should be useful for many different population genetic studies of the F. oxysporum species complex. Ethiopian F. oxysporum isolates, and F. oxysporum isolates obtained from other fungal collections, were characterized using AFLPs, SSRs and DNA sequence analyses. These studies resolved the isolates into three concordant groups that corresponded to the three clades of F. oxysporum previously designated for this species complex. Most of the Ethiopian isolates grouped in one of these three clades. This and the high degree of AFLP- and SSR-based genetic similarity observed among the Ethiopian isolates suggested that these isolates are largely clonal. In addition, a PCR-based diagnostic was developed to allow for the accurate differentiation between F. oxysporum and F. redolens which has proved to be difficult using morphological characters. The studies presented in this thesis contribute to a better understanding of Fusarium species commonly found in Ethiopia. Clearly, there is much that will still need to be done. The research presented in this thesis establishes a foundation for other studies and also provides a stimulus for further studies of these important fungi in Ethiopia. Copyright 2006, University of Pretoria. All rights reserved. The copyright in this work vests in the University of Pretoria. No part of this work may be reproduced or transmitted in any form or by any means, without the prior written permission of the University of Pretoria. Please cite as follows: Bogale, MA 2006, Molecular characterization of Fusarium isolates from Ethiopia, PhD thesis, University of Pretoria, Pretoria, viewed yymmdd < http://upetd.up.ac.za/thesis/available/etd-11072006-172206 / > D87/ag / Thesis (PhD)--University of Pretoria, 2007. / Genetics / Unrestricted

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Les études non destructives des processus physiologiques dans les produits agronomiques exigent des résolutions spatiales et temporelles de plus en plus élevées. L'imagerie par résonance magnétique nucléaire (RMN) est une technique totalement non-invasive qui permet d'accéder à plusieurs types de variables (architecture des tissus, variabilités spatiales de la composition, flux entrants et internes au cours de la croissance du fruit) plus difficilement quantifiables avec des méthodes destructives classiques. Un des enjeux majeur également réside dans la faculté de localiser spatialement ces transformations physiologiques et morphologiques dans les produits agronomiques. Les travaux de recherches réalisés dans le cadre de cette thèse ont pour objectif principal, la mise en œuvre d'une méthodologie de calcul et d'analyse quantitative en imagerie RMN appliquée à l'agronomie. L'implémentation, l'optimisation et la validation de la séquence FLASH combinée avec des agents de contraste efficaces en terme de relaxivité et bio-compatibles a permis d'une part, la cartographie des paramètres de relaxation et d'autre part, la quantification du transport de l'eau in vivo d'un système agronomique modèle au cours de sa croissance. Les nanoparticules de l'agent de contraste Gd3+[Fe(CN)6]3-/Mannitol ont été utilisées comme des marqueurs afin de localiser les flux hydriques dans le fruit. Le choix de la séquence d'imagerie FLASH a été motivé par la nécessite d'atteindre des résolutions temporelles suffisante pour suivre la dynamique des changements physiologiques liés au transport de l'eau dans ce type de matériau. La validation de la méthode de calcul du T1 menée sur le fantôme a révélé un bon accord par rapport aux T1 mesurés par relaxométrie. Nous avons également mis au point une procédure d'évaluation du rapport signal sur bruit et des incertitudes commises dans chaque voxel des images paramétriques M0 et T1. L'évaluation de ces incertitudes est un élément fondamental de cette analyse quantitative, afin d'assurer des interprétations fiables des images RMN. La segmentation des images nous a permis de localiser précisément les tissus où règne une forte activité cellulaire. Enfin, la modélisation compartimentale mis en oeuvre nous a permis de quantifier les paramètres cinétiques liés au transport de l'eau dans le fruit.Mots-clés: Imagerie RMN quantitative, paramètres intrinsèques, segmentation, modélisation compartimentale, agents de contraste, tissus végétaux / Non destructive studies of physiological processes in agronomic products require increasingly higher spatial and temporal resolutions. Nuclear Magnetic Resonance (NMR) imaging is a completely non-invasive technique providing access to several types of variables (tissue architecture, spatial variability of the composition, external and internal flow during fruit growth) more difficult to quantify with conventional destructive methods. One of major challenge lies in the ability to spatially localize the physiological and morphological changes in the agricultural products. The main objective of the research work in this thesis is to carry out a methodology in order to calculate and analyze quantitative NMR imaging applied to agronomy. The implementation, optimization and validation of the FLASH imaging sequence is performed in combination with innovative biocompatible contrast agents efficient in terms of relativity which allow to map in vivo relaxation parameters and then to explore water transportation in an agronomic model : the tomato during its growth. Nanoparticles of Gd3+[Fe(CN)6]3-/Mannitol contrast agents have been used as markers to localize the water flow in the fruit. The choice of the FLASH imaging sequence is motivated by the necessity to achieve sufficient high temporal resolution for monitoring the dynamics of physiological changes related to the water transport. The validation of the T1 calculation method performed on a phantom shows a good agreement compared to T1 measured by relaxometry. A systematic procedure for the estimation of the signal to noise ratio on the parametric images is also proposed which ensures a carefull determination of the intrinsic parameters of living tissues (M0 and T1) and their uncertainties. This step in the analysis ensures reliable interpretation of NMR images and permits image segmentation in order to precisely localize the tissues where there is a high cellular activity. Finally, the time dependance and the compartmental modeling allow to quantify the kinetic parameters associated with the water transport in the fruit.Keywords: Quantitative NMR imaging, intrinsic parameters, segmentation, compartmental modeling, contrast agents, plant tissues

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / This study evaluated the UV-C irradiation in tissues of rachis with and without berries, and berries with stems of Vitis vinifera and Vitis labrusca. In berries of cv. Isabella, the volatile compounds were analyzed on days 2, 5 and 10 of storage at 20 ºC and the effect of UV-C irradiation on the 5th day of storage. The experiments were based on four treatments of UV-C radiation (0, 1, 2 and 3 kJ m-2) with four cv. (Niagara white, Trebbiano, Cabernet sauvignon and Isabella) and evaluation times (three in berries and four rachis). The rachis with and without berries and the berries with stems remained stored at 20 ºC to evaluate physiological parameters. Ethylene production rate, respiratory rate and total polyphenols were evaluated in all tissues. Only in the rachis was determined by weight loss on days 1, 3 and 5, reducing sugars, non-reducing sugars on the 5th day. Berries and the juice from the cv. Isabella the 5th day of storage at 20 °C the total soluble solids (TSS), titratable acidity (TA), anthocyanins, total phenols, sugars and the ratio (TSS/ TA) were measured. In berries of cv. Isabella on days 2, 5 and 10 storage and berries irradiated with UV-C in amounts of 0, 1, 2 and 3 kJ m-2, volatiles were extracted by solid phase microextration. After desorption, the volatile compounds were separated by gas chromatography, identified by mass spectrometry and quantified by flame ionization detector. For the sensory analysis of the juice discriminatory tests for paired comparison of difference and test ranking were used. The physiological response of UV-C irradiation depends on the cv. and the irradiated tissue. Only the rachis of grape Vitis labrusca ‗Niagara white behaves climateric during storage in post-harvest and may reduce the self life of table grapes. This study found a possible hormetic dose of 3 kJ m-2 only for the cv. Isabella berry. 84 volatile compounds were detected over the days of storage. After 5 days storage, berries irradiated with UV-C 66 volatiles were found. And the dose of 3 kJ m-2 increased the concentration of esters and aldehydes, and 43,8% increase in the total polyphenols in berry. In the paired comparison test of difference, the judges showed greater intensity in the aroma of the juice from grapes irradiated. In the ranking test, the judges were unable to discriminate between different doses of irradiation, though differing dose control (0 kJ m-2) of the irradiated treatments. The analytical test enabled by gas chromatography to determine statistical differences between the different doses detected volatiles, especially at a dose of 3 kJ m-2, in which panelists found no difference. / Este trabalho avaliou a irradiação UV-C em tecidos vegetais de ráquis com e sem bagas, e bagas com pedúnculo de Vitis vinifera e Vitis labrusca. Nas bagas da cv. Isabel, foram analisados os compostos voláteis nos dias 2, 5 e 10 de armazenamento a 20 ºC e sobre o efeito da irradiação UV-C no 5º dia de armazenamento. Os experimentos basearam-se em quatro tratamentos de irradiação UV-C (0, 1, 2 e 3 kJ m-2), com quatro cv. (Niágara branca, Trebbiano, Isabel e Cabernet sauvignon) e tempos de avaliação (três nas bagas e quatro nas ráquis). As ráquis com e sem bagas e as bagas com pedúnculo permaneceram armazenadas a 20 ºC para avaliar parâmetros fisiológicos. Em todos os tecidos foram avaliadas a taxa de produção de etileno, taxa respiratória e polifenóis totais. Apenas na ráquis foi determinado a perda de peso nos dias 1, 3 e 5, além de açúcares redutores e não redutores no 5º dia. Nas bagas e no suco da cv. Isabel do 5º dia de armazenamento a 20 ºC foram mensurados os sólidos solúveis totais (SST), acidez total titulável (ATT), antocianinas totais, polifenóis totais, açúcares totais e a relação (SST/ATT). Nas bagas da cv. Isabel dos dias 2, 5 e 10 de armazenamento e nas bagas irradiadas com UV-C nas doses de 0, 1, 2 e 3 kJ m-2, foram extraídos os compostos voláteis por microextração em fase sólida. Após a dessorção, os compostos voláteis foram separados em cromatógrafo a gás, identificados pelo espectrômetro de massas e quantificados por detector de ionização em chamas. Para a análise sensorial do suco, foram utilizados os testes discriminativos de comparação pareada de diferença e teste de ordenação. A resposta fisiológica da irradiação UV-C depende da cv. e do tecido irradiado. Apenas a ráquis da uva Vitis labrusca ‗Niágara branca se comporta de forma climatérica durante o armazenamento na pós-colheita, podendo reduzir a vida de prateleira da uva de mesa. Neste estudo foi encontrada uma possível dose hormética de 3 kJ m-2 somente para a baga da cv. Isabel. Foram detectados 84 compostos voláteis ao longo dos dias de armazenamento. No 5º dia de armazenamento, das bagas irradiadas com UV-C, foram encontrados 66 compostos voláteis, sendo que a dose de 3 kJ m-2 aumentou a concentração de ésteres e aldeídos, além de elevar em 43,8% os polifenóis totais na baga. No teste de comparação pareada de diferença, os provadores indicaram maior intensidade no aroma do suco proveniente de uvas irradiadas. No teste de ordenação, os julgadores não conseguiram discriminar as diferentes doses de irradiação, mas diferenciaram a dose controle (0 kJ m-2) dos tratamentos irradiados. O teste analítico por cromatografia gasosa permitiu determinar diferença estatística entre as doses nos diferentes compostos voláteis detectados, principalmente na dose de 3 kJ m-2, na qual os julgadores não encontraram diferença.

Die Vorverarbeitung und Analyse von spektrometrischen und spektroskopischen Daten von Pflanzengewebe sind in den unterschiedlichsten Forschungsbereichen wie der Pflanzenbiologie, Agrarwissenschaften und Klimaforschung von großer Bedeutung. Der Schwerpunkt dieser Arbeit liegt auf der optimierten Nutzung von Daten von Pflanzengeweben, insbesondere der Daten gewonnen durch Matrix–Assistierte Laser–Desorption–Ionisierung Massenspektrometrie, Raman-Spektroskopie und Fourier-Transform-Infrarotspektroskopie. Die Klassifizierungsfähigkeit mit diesen Methoden wird insbesondere nach Kombination der Daten untereinander und mit zusätzlichen chemischen und biologischen Informationen verglichen. Die diskutierten Beispiele befassen sich mit der Untersuchung und Einordnung innerhalb einer bestimmten Pflanzenart, beispielsweise der Unterscheidung von Proben aus unterschiedlichen Populationen, Wachstumsbedingungen oder Gewebeunterstrukturen. Die Daten wurden mit sowohl mit explorativen Werkzeugen wie der Hauptkomponentenanalyse und der hierarchischen Clusteranalyse, als auch mit Methoden des maschinellen Lernens wie die Diskriminanzanalyse oder künstliche neuronale Netzwerke umfassten. Konkret zeigen die Ergebnisse, dass die Kombination der Methoden mit zusätzlichen pflanzenbezogenen Informationen in einer Konsensus-Hauptkomponentenanalyse zu einer umfassenden Charakterisierung der Proben führt. Es werden verschiedene Strategien zur Datenvorbehandlung diskutiert, um nicht relevante spektrale Information zu reduzieren, z.B. aus Karten von Pflanzengeweben oder eingebetteten Pollenkörnern. Die Ergebnisse dieser Arbeit weisen auf die Relevanz der gezielten Nutzung spektrometrischer und spektroskopischer Daten hin und lassen sich nicht nur auf pflanzenbezogene Themen, sondern auch auf andere analytische Klassifizierungsprobleme übertragen. / The pre-processing and analysis of spectrometric and spectroscopic data of plant tissue are important in a wide variety of research areas, such as plant biology, agricultural science, and climate research. The focus of the thesis is the optimized utilization of data from plant tissues, which includes data from Matrix-Assisted-Laser Desorption/Ionization time of flight mass spectrometry, Raman spectroscopy, and Fourier transform infrared spectroscopy. The ability to attain a classification using these methods is compared, in particular after combination of the data with each other and with additional chemical and biological information. The discussed examples are concerned with the investigation and classification within a particular plant species, such as the distinction of samples from different populations, growth conditions, or tissue substructures. The data were analyzed by exploratory tools such as principal component analysis and hierarchical cluster analysis, as well as by predictive tools that included partial least square-discriminant analysis and machine learning approaches. Specifically, the results show that combination of the methods with additional plant-related information in a consensus principal component analysis leads to a comprehensive characterization of the samples. Different data pre-treatment strategies are discussed to reduce non-relevant spectral information, e.g., from maps of plant tissues or embedded pollen grains. The results in this work indicate the relevance of the targeted utilization of spectrometric and spectroscopic data and could be applied not only to plant-related topics but also to other analytical classification problems.