Magistrantūros studijų darbu siekta nustatyti Leptosphaeria maculans ir L. biglobosa rūšių paplitimą ir pasiskirstymą ant įvairių rūšių bastutinių šeimos augalų (Brassica napus var. biennis, Brassica napus var. annua, Brassica oleracea var. capitata, Brassica oleracea var. italica) bei grybo surinkto iš skirtingų augalų rūšių augimo specifiką in vitro sąlygomis. / The master work was aimed to identify the occurrence and distribution of Leptosphaeria maculans and Leptosphaeria biglobosa species on the plants of various Brassica species (Brassica napus var. biennis, Brassica napus var. annua, Brassica oleracea var. capitata, Brassica oleracea var. italica) determine growth specificity of fungi on agar media, collected from different plant species, in in vitro conditions.

An investigation was made into the isolation and screening of highly cellulolytic anaerobic fungi and their production of cellulolytic enzymes using immobilised rhizomycelia. A total of 46 anaerobic fungi were isolated on cellulosic substrates from ruminant and non-ruminant herbivores. Primary screening of these isolates was performed using dye release from cellulose-azure which qualitatively detected cellulolytic activity. Twelve isolates were chosen on the basis of their maximum solubilisation rates of the labelled cellulose and then subjected to secondary screening which involved the quantification of enzyme activity. The enzyme mixtures were characterised by carboxymethylcellulase, xylanase, B-glucosidase, B-xylosidase and cellobiase assays, measured by the production of either reducing sugars, p-nitrophenol or glucose. All strains produced a number of enzymes that allowed them to hydrolyse straw and highest enzyme activity was measured in static cultures grown on 0.5% straw. A monocentric isolate, Piromyces strain KSX1 from a red kangaroo, and a cattle polycentric isolate, Orpinomyces strain 478P1, were selected for study of cellulolytic enzyme production on the basis of high fibre digestion capability and amenability toward encapsulation. The immobilised polycentric strain proved to be operationally superior to strain KSX1 as strain 478P1 did not produce any viable growth in the culture liquor. Studies into single batch cultures of free cells of strains KSX1 and 478P1 revealed that the maximum specific rate of B-glucosidase production occurred concomitantly with maximum specific growth rate suggesting that the immobilised fungus must grow for continuous enzyme production to occur. Although the physiology of cellulase synthesis in strains KSX1 and 478P1 was found to be growth-associated, immobilisation of the fungus offered the advantage of the repeat-batch use of cells with the accumulation of extracellular enzymes after each batch. Thus, operational gains were the key issues in assessing the potential application of immobilised anaerobic fungi in the production of cellulolytic enzymes. The repeat-batch system was operationally more efficient than the free cell batch cultures because immobilisation removed the need of reculturing the cells for every single batch. / Doctor of Philosophy (PhD)

Background: Pancreatic cancer is one of the leading causes of cancer-related death. Using DNA gene expression analysis based on a custom made Affymetrix cancer array, we investigated the expression pattern of both primary and established pancreatic carcinoma cell lines. Methods: We analyzed the gene expression of 5 established pancreatic cancer cell lines (AsPC-1, BxPC-3, Capan-1, Capan-2 and HPAF II) and 5 primary isolates, 1 of them derived from benign pancreatic duct cells. Results: Out of 1,540 genes which were expressed in at least 3 experiments, we found 122 genes upregulated and 18 downregulated in tumor cell lines compared to benign cells with a fold change > 3. Several of the upregulated genes (like Prefoldin 5, ADAM9 and E-cadherin) have been associated with pancreatic cancer before. The other differentially regulated genes, however, play a so far unknown role in the course of human pancreatic carcinoma. By means of immunohistochemistry we could show that thymosin [β-10 (TMSB10), upregulated in tumor cell lines, is expressed in human pancreatic carcinoma, but not in non-neoplastic pancreatic tissue, suggesting a role for TMSB10 in the carcinogenesis of pancreatic carcinoma. Conclusion: Using gene expression profiling of pancreatic cell lines we were able to identify genes differentially expressed in pancreatic adenocarcinoma, which might contribute to pancreatic cancer development. / Dieser Beitrag ist mit Zustimmung des Rechteinhabers aufgrund einer (DFG-geförderten) Allianz- bzw. Nationallizenz frei zugänglich.

The cultivation of flowers is, in Brazil, very recent and little known species have a high ornamental potential, Cartamus is such a species and there are cultivars for the production of oil and as well as for ornamental pourposes. Being a recent crop in Brazil, little is known about its associated diseases, specially those that affect the seeds and the soil borne, such as white mold that causes great losses to several crops. Six experiments were conducted with the objectives of testing the reaction of the crop to isolates of Sclerotinia sclerotiorum, the biocontrol of S. sclerotiorum and of pathogens associated to the seeds, and the effect of the biocontrollers on cartamus plant growth. Isolates of the pathogen from chrisantemum, lettuce, soybean, and carrots, as well as isolates of Trichoderma sp. (ETSR 20 e TC 1.15) and comercial products of Trichoderma spp. (Agrotrich and Trichodel®) were used. In the reaction test, the cartamus crop was more severely attacked by the isolate obtained from lettuce, which when incorporated to commercial substrate did not promote the development of the disease in plant, but reduced plant growth. Trichodel® was the product that promoted the highest growth of plants when incorporated to the substrate, even in the absence of the pathogen. The product more efficient in controlling seed pathogens was Agrotrich. In the growth of seedlings, the isolates ETSR20 and TC1.15 were the best, when applied to the seeds and the latter promoted the best emergency of seeds in commercial substrate. Therefore, the Trichoderma based products can be used in the control of seed pathogens and growth promotion of cartamus plants as seed or substrate treatment. There are differences in disease severity on cartamus among isolates of S. sclerotiorum from different crops and its presence reduces the crop s growth, even in the absence of visible symptoms / A floricultura, no Brasil, é relativamente recente, existindo espécies pouco conhecidas com alto potencial ornamental. O cártamo é uma delas e possui cultivares tanto para a produção de óleo quanto para ornamentação. Por ser uma cultura recente no Brasil, pouco se conhece sobre as doenças associadas a ela, especialmente as que afetam as sementes e as veiculadas pelo solo, como o mofo branco, que causa grandes prejuízos a diversas culturas. Seis experimentos foram conduzidos com o objetivo de testar a reação da cultura a isolados de Sclerotinia sclerotiorum, o biocontrole de patógenos associados às sementes e de Sclerotinia sclerotiorum e o efeito dos biocontroladores no crescimento de plantas de cártamo. Foram utilizados isolados de Sclerotinia sclerotiorum das culturas do crisântemo, alface, soja e cenoura, e isolados de Trichoderma spp. (ETSR 20 e TC 1.15) e formulados comerciais à base de Trichoderma spp. (Agrotrich e Trichodel®). No teste de reação, a cultura do cártamo foi mais severamente atacada pelo isolado proveniente da alface, o qual, quando incorporado ao substrato comercial, não promoveu o desenvolvimento da doença nas plantas, porém prejudicou seu crescimento. Trichodel® foi o produto que maior crescimento proporcionou às plantas quando incorporado ao substrato, mesmo sem a presença do patógeno. O produto mais eficiente no controle dos patógenos das sementes foi o formulado comercial Agrotrich. No crescimento de plântulas, os isolados ETSR 20 e TC 1.15 se sobressaíram quando aplicados às sementes e este obteve melhor resultado na emergência de plântulas em substrato. Assim, os produtos a base de Trichoderma são viáveis para controle de patógenos de sementes e crescimento de plantas tanto no tratamento de sementes como do substrato. Existem diferenças na severidade da doença em cártamo entre isolados de Sclerotinia sclerotiorum oriundos de diferentes culturas e sua presença reduz o crescimento da cultura, mesmo na ausência de sintomas visíveis

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No. of bitstreams: 1 Tese - Arlena Maria Guimarães Gato.pdf: 2067444 bytes, checksum: a858fc3c2c47508ee175666940ad0251 (MD5) Previous issue date: 2009-12-10 / SUFRAMA - Superintendência da Zona Franca de Manaus / The flowers and ornamental plants cultivation stands out as an important agronomical activity. But despite the economical and market potential of these regional native species: helicônia, bastão, sorvetão and other plants, is necessary more basic studies about the use of advanced techniques in getting propagated material with quality assurance and good phytosanitary aspect. The objective of this study was to obtain plantlets from floral apices of Heliconia rauliniana and embryos rescue of H. marginata, identifies, inoculation of bacterial isolates and evaluate the effect of these microorganisms in the development of micropropagated plants. Established the plants micropropagation process, it was isolated from matrices root, bacterial isolates and, according to the classification criteria of activity levels of nitrogen biological fixation, phosphate solubilisation and auxin production, it was selected six Heliconia rauliniana isolates and eight of Heliconia marginata. The first experiment were inoculated in in vitro plants of H. rauliniana, the B4, B5, B8, B13, B16 and B18 bacterial isolates and on H. marginata micropropagated plants, the B1, B2, B4, B6, B7, B10, B12, B14 and B16 isolates and, then transferred to plastic boxes containing sterilized substrate Plantmax (Horticulture) and maintained under greenhouse. After 60 days of inoculation, it was realized the evaluation of root growth promotion, selecting the three isolates that showed the best results: B18, B5 and B13 (H. rauliniana) and B7, B6 and B10 (H. marginata). In the second experiment we used the B18, B5, B13 and B6, B7, B10 selected in the first experiment and inoculated in 60 plants, distributed on five treatments: control, B18, B5 and B13, cocktail x plant and control, B6, B7, B10, cocktail x plant with 12 plants per treatment. The results presented after 60 days, don’t were significant on the level of 0.05% and one good faith coefficient of 95% (ANOVA) for root growth promotion, number of leaves, plant height, fresh weight and dry weight. The plants survive was 83.3%. For microorganisms identify, it was used the analysis of the fragment about 800 bp of the 16S rDNA (Primer 27f). The sequencial analysis via Blast showed three bacterial groups: Burkholderia, Ralstonia e Enterobacteriaceae (Pantoea/Erwinia). / O cultivo de flores e plantas ornamentais tropicais destaca-se como importante atividade agrícola. Mas, apesar das potencialidades econômicas e de mercado dessas espécies nativas da região: helicônia, bastão, sorvetão e outras, são necessários mais estudos básicos sobre a utilização de técnicas avançadas na obtenção de material de propagação com garantia de qualidade e bom aspecto fitossanitário. O objetivo deste trabalho foi obter mudas micropropagadas a partir de ápice floral de Heliconia rauliniana e de resgates de embriões de H. marginata, identificação, inoculação dos isolados de bactérias e avaliar o efeito desses microrganismos no desenvolvimento das plantas micropropagadas. Estabelecido o processo de micropropagação das plantas foram isolados das raízes das matrizes, isolados bacterianos e, de acordo com os critérios de classificação dos níveis de atividades em fixação biológica de nitrogênio, solubilização de fosfato e produção de auxina, foram selecionados seis isolados da Heliconia rauliniana e oito da Heliconia marginata. No primeiro experimento, foram inoculados nas plantas micropropagadas de H. rauliniana, os isolados bacterianos B4; B5 (Enterobacter); B8; B13 (Ralstonia); B16; B18 (Enterobacter) e nas plantas micropropagadas de H. marginata, os isolados B1; B2; B4; B6 (Enterobacter); B7 (Enterobacter); B10 (Burkholderia); B12; B14 e; B16 e, posteriormente transferidas para caixas plásticas contendo substrato esterilizado PlantMax (Horticultura) e mantidas em casa de vegetação. Após 60 dias de inoculação foi realizada a avaliação de promoção de crescimento de raiz, selecionando-se os três isolados que apresentaram os melhores resultados por espécie: Heliconia rauliniana, Enterobacteriaceae (Pantoea/Erwinia), B18 e B5 Enterobacteriaceae (Pantoea/Erwinia e B13 (Ralstonia) e Heliconia marginata B7 Entereobacteriaceae (Pantoea/Erwinia), B6 Entereobacteriaceae (Pantoea/Erwinia) e B10 Burkholderia. No segundo experimento foram utilizados os isolados bacterianos B18 Enterobacteriaceae (Pantoea/Erwinia): B5 Enterobacteriaceae (Pantoea/Erwinia, B13 Ralstonia e B6, Entereobacteriaceae (Pantoea/Erwinia), B7, Entereobacteriaceae (Pantoea/Erwinia), B10, Burkholdeia selecionados no primeiro experimento e inoculados em 60 plantas, distribuídas em cinco tratamentos: controle, B18 Entereobacteriaceae (Pantoea/Erwinia: B5 Entereobacteriaceae (Pantoea/Erwinia e B13 Ralstonia, coquetel x planta e controle, B6 Entereobacteriaceae (Pantoea/Erwinia), B7 Entereobacteriaceae (Pantoea/Erwinia), B10 (Burkholderia), coquetel x planta com 12 plantas por tratamento. Os resultados apresentados após 60 dias, não foram significativos a nível de 0,05 % e um coeficiente de confiança de 95 % (A NOVA) para promoção de crescimento de raiz, número de folhas, altura de planta, peso fresco e peso seco. A sobrevivência das plantas foi de 83,3 %. Para identificação dos microrganismos foi usado o seqüenciamento de fragmento de aproximadamente 800 pb do gene 16S rDNA (“primer 27f). A análise das sequencias via Blast mostrou três grupos de bactéria: Burkholderia, Ralstonia e Enterobacteriaceae (Pantoea/Erwinia).

This diploma thesis deals with the use of various types of bran as a by-product in the milling of cereals. Mills create a huge amount of this material per year. The most common way of processing bran is mostly incineration and to a lesser extent it is used as feed for livestock. Depending on the type of cereal, bran contains 10-20% of protein, which disappears from the food chain due to combustion. Within the framework of sustainability and valorisation of waste, which has recently been largely discussed, great emphasis is placed on waste minimization whether in the field of its production or further processing. Due to the relatively high protein content, bran appears to be a suitable starting material to produce protein supplements. Proteins can be extracted from bran based on their different solubility at different pH. In the alkaline method, the proteins are first dissolved in an alkaline pH and then precipitated in an acidic medium. Lyophilization is followed by characterization of the extract in terms of yield, protein content, moisture, amino acid profile and digestibility. The highest yield was obtained with the oat bran isolate (13,5 ± 0,6 g of isolate per 100 g of bran). In terms of protein content, the best protein isolate was also obtained from oat bran (95,2 ± 0,4% protein in the isolate). Another determination was the analysis of the amino acid profile, in which a high content of arginine was found in all analyzed protein isolates from bran. Determination of digestibility showed very good digestibility of all produced protein extracts from bran.

Streptomycetes, primary soil saprophytic microorganisms are at the center of interest in many research groups, mainly because of their ability to produce a wide range of biologically active substances useful in medicine, biotechnology and agriculture. The marginal, and little explored areas are the interactions that streptomycetes create with humans. Recent metagenomic studies have shown that streptomycetes colonize the skin, the respiratory and possibly the urogenital tract of humans. In addition to apparent pathogens such as S. somaliensis and S. sudanensis, the clinical impact of these streptomycetes on human health is unknown. For this reason, a unique collection of non-pathogenic streptomycetes isolated from human clinical specimens was developed. The isolates were collected by the National Reference Laboratory for Pathogenic Actinomycetes in Trutnov, Czech Republic. On the basis of pilot studies, an isolate labeled TR42 was selected from the Trutnov collection, showing a very broad spectrum of biological activities. This strain was isolated from sputum from a patient with unknown respiratory diagnosis. The TR42 strain exhibits considerable biotechnological potential and after following a thorough study, it could be a source of new biologically active substances with pharmaceutically...

<p>Regulatory interactions evolve to incorporate new genomic material and contribute to bacterial diversity. These regulatory interactions are flexible and likely provide bacteria with a means of rapid environmental adaptation. In this thesis, the RpoS regulon is used as a model system to investigate the hypothesis that regulon composition and expression are modified according to environmental pressures. Several novel findings are presented, namely the distribution of RpoS homologs in bacteria, the flexibility of the RpoS regulon, and the effect of diverse environmental pressures on RpoS regulon expression. Based on phylogenetic and reciprocal best hits analyses, RpoS was determined to be conserved in gamma-, beta-, and delta-proteobacteria, likely because it confers a selective advantage in many bacterial niches. Regulon composition, however, was highly flexible. Even between species of the same class, <em>Escherichia coli</em> and <em>Pseudomonas aeruginosa</em>, only 12 of 50 orthologs were regulated in common by RpoS. RpoS regulon flexibility may thus be the result of adaptation to different bacterial habitats. Indeed, mutations in <em>rpoS</em> and differential regulon expression could be identified among environmental <em>E. coli</em> isolates collected from diverse sources. Among environmental <em>E. coli</em> isolates, RpoS mutant frequency was found to be 0.3%, and activity of KatE, a prototypical RpoS regulon member, was undetectable in some isolates despite the presence of functional RpoS. Modulated RpoS regulon expression among environmental <em>E. coli</em> isolates is consistent with environment as a key factor shaping regulatory interactions. Regulon flexibility was similarly apparent in oxidative stress regulons, OxyR and SoxRS, of <em>E. coli</em>. SoxRS regulon function is weakly conserved, possibly due to low selective pressure for a superoxide stress response regulon in some bacterial species. Environment, therefore, is a crucial element that defines the dynamics of regulatory networks.</p> / Doctor of Philosophy (PhD)

Aeromonas hydrophila is distributed widely in nature and is responsible for an array of human infections. Several studies on the isolation and characterisation of the organism abound. Although there are reports on the antibiotic resistance profiles of the organism, these reports have not been updated in Limpopo province, South Africa despite the established fact that antibiograms vary with time and geographical area. Antibiotic resistance and pathogenesis of an organism are dependent on a host of factors such as the production of extended spectrum beta-lactamases and the genetic profiles such as the genes coding for resistance and possession of integrons and how these characteristics overach with the phylogenetic inter-relatedness of isolates from different sources. In spite of the aforementioned concerns on the efficacy of antibiotics due to the acquisition or endowment of microorganisms with intrinsic and extrinsic factors , which enhances resistance to antibiotics , medicinal plants are reportedly offering promise as alternative sources of efficacious management of infections. Medicinal plants are employed by traditional healers in the management of infections in developing countries especially in Africa. However, the antimicrobial activities of medicinal plants against Aeromonas hydrophila have received only a cursory attention. In an endeavour to undertake a comprehensive study on the isolation, characterisation, antibiograms, activities of medicinal plants as well as the genetic profiles, including phylogenetics relatedness of Aeromonas isolates from different sources, stool and water samples were collected over a two year period from designated places in Limpopo Province and analysed using standard techniques applicable to the constituent research activity. The research findings are presented in six chapters as presented hereunder. The first chapter focussed on the literature review of the organism and reflects areas such as the morphology, laboratory diagnosis, clinical manifestations, pathogenesis, antimicrobial susceptibility profiles, antibacterial activities of medicinal plants as well as the genetic aspects of Aeromonas hydrophila. / Environmental Science / D.Phil. (Environmental Science)

Doctor of Philosophy / Department of Plant Pathology / Megan Kennelly / Large patch, caused by the fungus Rhizoctonia solani anastomosis group (AG) 2-2 LP, is the most common and severe disease of zoysiagrass (Zoysia spp). Despite the importance of this disease, few studies have examined pathogen biology, cultivar susceptibility, cultural controls, and chemical controls. The objectives of this dissertation were: (1) Characterize large patch isolates based on anastomosis pairing, in-vitro mycelial growth rates, nuclear counts, virulence, PCR, and amplified fragment length polymorphism (AFLP); (2) Determine the effects of cultivation (aerification, verticutting, and sand topdressing) on disease severity; (3) Evaluate different fall and spring applications of the fungicides flutolanil, azoxystrobin, and triticonazole; (4) Evaluate the susceptibility of fifteen new zoysiagrass germplasm lines from parental crosses including Z. japonica, Z. matrella, and Z. pacifica. All the R. solani isolates from large patch-infected zoysiagrass from Kansas belonged to AG 2-2 LP. Variations were observed among the isolates in their average number of nuclei per cell, mycelial growth rates and virulence. There was also variation in the amplified fragment length polymorphism (AFLP) DNA fingerprints, suggesting possible underlying genetic differences of biological significance among members of AG 2-2 LP. Cultivation did not affect soil moisture or temperature. Cultivation also did not reduce patch sizes, nor influence turf recovery rate from large patch. From 2009 to 2011, spring and fall N fertility was consistently associated with lower percentages of diseased turf in both cultivated and non-cultivated plots at Manhattan and Haysville. In general, two fall applications of fungicide did not reduce disease compared to one fall application. Fungicides applied in the fall when thatch temperatures ranged from 17.8oC to 23.2oC reduced disease compared to untreated controls. Early spring applications reduced disease compared to later spring applications. In germplasm screening studies, all progeny had similar disease levels compared to Meyer in the growth chamber, but only 6 consistently had disease levels as low as Meyer in the field. Growth chamber results did not correlate to field results.