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Williams, Katherine. "Genetic manipulation of fungal secondary metabolism." Thesis, University of Bristol, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.535469.
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Delsol, Anne Aline Germaine. "Microbial 7-hydroxylation of the steroid lithocholic acid : a novel approach to produce bile acids for gallstone therapy." Thesis, University of Exeter, 1999. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.297640.
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Bakker, Walid Ismail Mohammed Mohammed. "Overexpression of secondary metabolism genes from Magnaporthe grisea and Beauveria bassiana speciality : fungal biotechnology." Thesis, University of Bristol, 2011. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.544416.
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Pfannenstiel, Brandon T., Xixi Zhao, Jennifer Wortman, Philipp Wiemann, Kurt Throckmorton, Joseph E. Spraker, Alexandra A. Soukup, et al. "Revitalization of a Forward Genetic Screen Identifies Three New Regulators of Fungal Secondary Metabolism in the Genus Aspergillus." AMER SOC MICROBIOLOGY, 2017. http://hdl.handle.net/10150/626452.
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The study of aflatoxin in Aspergillus spp. has garnered the attention of many researchers due to aflatoxin's carcinogenic properties and frequency as a food and feed contaminant. Significant progress has been made by utilizing the model organism Aspergillus nidulans to characterize the regulation of sterigmatocystin (ST), the penultimate precursor of aflatoxin. A previous forward genetic screen identified 23 A. nidulans mutants involved in regulating ST production. Six mutants were characterized from this screen using classical mapping (five mutations in mcsA) and complementation with a cosmid library (one mutation in laeA). The remaining mutants were backcrossed and sequenced using Illumina and Ion Torrent sequencing platforms. All but one mutant contained one or more sequence variants in predicted open reading frames. Deletion of these genes resulted in identification of mutant alleles responsible for the loss of ST production in 12 of the 17 remaining mutants. Eight of these mutations were in genes already known to affect ST synthesis (laeA, mcsA, fluG, and stcA), while the remaining four mutations (in laeB, sntB, and hamI) were in previously uncharacterized genes not known to be involved in ST production. Deletion of laeB, sntB, and hamI in A. flavus results in loss of aflatoxin production, confirming that these regulators are conserved in the aflatoxigenic aspergilli. This report highlights the multifaceted regulatory mechanisms governing secondary metabolism in Aspergillus. Additionally, these data contribute to the increasing number of studies showing that forward genetic screens of fungi coupled with whole-genome resequencing is a robust and cost-effective technique. IMPORTANCE In a postgenomic world, reverse genetic approaches have displaced their forward genetic counterparts. The techniques used in forward genetics to identify loci of interest were typically very cumbersome and time-consuming, relying on Mendelian traits in model organisms. The current work was pursued not only to identify alleles involved in regulation of secondary metabolism but also to demonstrate a return to forward genetics to track phenotypes and to discover genetic pathways that could not be predicted through a reverse genetics approach. While identification of mutant alleles from whole-genome sequencing has been done before, here we illustrate the possibility of coupling this strategy with a genetic screen to identify multiple alleles of interest. Sequencing of classically derived mutants revealed several uncharacterized genes, which represent novel pathways to regulate and control the biosynthesis of sterigmatocystin and of aflatoxin, a societally and medically important mycotoxin.
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Waldenmaier, Hans Eugene. "Bioluminescência fúngica: papel ecológico, purificação e clonagem de enzimas." Universidade de São Paulo, 2016. http://www.teses.usp.br/teses/disponiveis/46/46131/tde-14072017-145527/.
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Esta tese de doutorado descreve os estudos realizados para elucidar a biologia molecular da bioluminescência fúngica e sua relevância ecológica na natureza. A recente descoberta de que a luciferina fúngica é a 3-hidroxihispidina permitiu a caracterização do metabolismo secundário da fenilalanina nos genomas recém-sequenciados e transcriptomas de micélios das espécies luminescentes Panellus stipticus e Neonothopanus gardneri. Adicionalmente os genomas e transcriptomas de variedades não luminescente de P. stipticus e Lentinula edodes serviram como respectivos controles. Em geral, os genes envolvidos no metabolismo secundário da fenilalanina em amostras luminescentes tinham expressão igual ou superior àquela de espécies não luminescentes. Um agrupamento de genes relacionados com a biossíntese de fenilalanina foi encontrado em ambos os genomas luminescentes e não luminescentes de P. stipticus. A abundância de genes transcritos neste agrupamento foi semelhante para as espécies luminescentes e não luminescentes de P. stipticus, mas a policetídeo sintase tipo I em P. stipticus não luminescentes foi significativamente sub-regulada. Não foi encontrado agrupamento semelhante nos genomas de N. gardneri e L. edodes, sendo que os correspondentes homólogos estavam espalhados em diferentes loci. Extratos de fungos podem ser preparados in vitro, com a adição de 3-hidroxihispidina para produzir luz verde em abundância. A preparação de extratos proteicos de luciferase foi melhorada e a estrutura da luciferase, parcialmente purificada, foi investigada por espectrometria de massas. A presença de luciferase nos géis de purificação foi revelada usando-se luciferina e molécula similares à luciferina advindas de extratos de plantas. O nicho ecológico nas vizinhas de cogumelos bioluminescentes foi investigado de duas maneiras, armadilhas adesivas com cogumelos artificiais de acrílico, iluminados com luz LED verde e através da observação direta de cogumelos bioluminescentes com fotografia no infravermelho com lapso de tempo. Os estudos ecológicos foram conduzidos nos biomas da Mata Atlântica e da Mata dos Cocais, no Brasil. Baratas, aranhas, tesourinhas, grilo e vagalumes tec-tecs foram os animais mais comuns que interagiram com os cogumelos. Todos estes animais podem agir como dispersores de propágulos e, em alguns casos, como defensores dos cogumelos.This PhD thesis describes the studies performed to elucidate the molecular biology of fungal bioluminescence and the ecological significance of the trait in the wild. The recent discovery that the fungal luciferin is 3-hydroxyhispidin has allowed for the characterization of phenylalanine secondary metabolism in the newly sequenced genomes and mycelium transcriptomes of luminescent Panellus stipticus and Neonothopanus gardneri, additionally the genomes and transcriptomes of a non-luminescent variety of P. stipticus and Lentinula edodes served as respective controls. In general the genes involved in phenylalanine secondary metabolism had greater or equal expression in luminescent samples than non luminescent. A cluster of genes related to the secondary metabolism of phenylalanine was found in both luminescent and non luminescent P. stipticus genomes. Transcript abundance of genes in this cluster was similar in both luminescent and non-luminescent Panellus stipticus, but the type I polyketide synthase in non luminescent Panellus stipticus was significantly down regulated. A similar gene cluster in the N. gardneri and L. edodes genomes was absent with corresponding homologues scattered at different genomic loci. Cell free fungal extracts can be combined in vitro with the addition of 3-hydroxyhispidin to produce abundant green light. Preparation of proteinaceous luciferase extracts was improved and partially purified luciferase samples were investigated by mass spectrometry. The presence of luciferase in the separation gel was also evidenced by using luciferin and luciferin-like molecules from plant extracts. The ecological niche surrounding bioluminescent mushrooms was investigated through two main means, glue traps with acrylic mushroom facsimiles that were internally illuminated with green LED lights and direct observation of bioluminescent mushrooms with infrared time lapse photography. Ecological studies were performed in the Atlantic rainforest (Mata Atlântica) and transitional Coconut Palm forest (Mata dos Cocais) biomes of Brazil. Cockroaches, spiders, earwigs, crickets, and luminescent click beetles were the most common animal interacting with mushrooms. All of these animals may be acting as fungal propagule dispersers and in some cases defense of the mushroom.
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Meister, Cindy [Verfasser], Gerhard H. [Akademischer Betreuer] Braus, Gerhard H. [Gutachter] Braus, Kai [Gutachter] Tittmann, and Achim [Gutachter] Dickmanns. "Interplay of the COP9 signalosome deneddylase and the UspA deubiquitinase to coordinate fungal development and secondary metabolism / Cindy Meister ; Gutachter: Gerhard H. Braus, Kai Tittmann, Achim Dickmanns ; Betreuer: Gerhard H. Braus." Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2019. http://d-nb.info/1187520330/34.
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Maertens, Jeroen Moritz. "Reconstruction of fungal secondary metabolite biosynthetic pathways in Aspergillus oryzae." Thesis, University of Bristol, 2016. https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.738201.
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Turner, Adrian Simon. "An investigation into the switch between primary and secondary metabolism in Cephalosporium acremonium." Thesis, University of Liverpool, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.240785.
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Kim, Kwang Hyung. "Functional Analysis of Secondary Metabolite Biosynthesis-Related Genes in Alternaria brassicicola." Diss., Virginia Tech, 2009. http://hdl.handle.net/10919/39452.
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Alternaria brassicicola is a necrotrophic pathogen that causes black spot disease on virtually all cultivated Brassicas, A. brassicicola is renowned for its ability to prodigiously produce secondary metabolites. To test the hypothesis that secondary metabolites produced by A. brassicicola contribute to pathogenicity, we identified seven nonribosomal peptide synthetases (NPSs) and 10 polyketide synthases (PKSs) in the A. brassicicola genome. The phenotype resulting from knockout mutations of each PKS and NPS gene was investigated with an emphasis on discovery of fungal virulence factors. A highly efficient gene disruption method using a short linear double stranded DNA construct with minimal elements was developed, optimized, and used to functionally disrupt all NPS and PKS genes in A. brassicicola. Three NPS and two PKS genes, and one NPS-like gene appeared to be virulence factors based upon reduced lesion development of each mutant on inoculated green cabbage and Arabidopsis compared with the wild-type strain. Furthermore some of the KO mutants exhibited developmental phenotypic changes in pigmentation and conidiogenesis. To further characterize the roles of several genes of interest in A. brassicicola development and pathogenesis, the genes AbNPS2, AbPKS9, and NPS-like tmpL were selected for in-depth functional analysis. We provide substantial evidence that the AbNPS2-associated metabolite is involved in conidial cell wall construction, possibly as an anchor connecting two cell wall layers. We also characterized a biosynthetic gene cluster harboring the AbPKS9 gene and demonstrated that this cluster is responsible for the biosynthesis of depudecin, an inhibitor of histone deacetylases and a minor virulence factor. Finally, we demonstrated that a NPS-like protein named TmpL is involved in a filamentous fungi-specific mechanism for regulating levels of intracellular reactive oxygen species during conidiation and pathogenesis in both plant and animal pathogenic fungi. Collectively our results indicate that small molecule nonribosomal peptides and polyketides in A. brassicicola play diverse, but also fundamental, roles in fungal development and pathogenesis.Ph. D.
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Kutil, Brandi Lynn. "The evolution of LOL, the secondary metabolite gene cluster for insecticidal loline alkaloids in fungal endophytes of grasses." Thesis, [College Station, Tex. : Texas A&M University, 2006. http://hdl.handle.net/1969.1/ETD-TAMU-1122.
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Vaughan, Martha Marie. "Molecular and Functional Characterization of Terpene Chemical Defense in Arabidopsis Roots in Interaction with the Herbivore Bradysia spp. (fungus gnat)." Diss., Virginia Tech, 2010. http://hdl.handle.net/10919/77974.
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Roots and leaves are integrated structural elements that together sustain plant growth and development. Insect herbivores pose a constant threat to both above- and belowground plant tissues. To ward off herbivorous insects, plants have developed different strategies such as direct and indirect chemical defense mechanisms. Research has primarily focused on visible aboveground interactions between plants and herbivores. Root-feeding insects, although often overlooked, play a major role in inducing physical and physiological changes in plants. However, little is known about how plants deploy chemical defense against root herbivores.
We have developed an Arabidopsis aeroponic culture system based on clay granulate, which provides access to root tissue and accommodates subterranean insect herbivores. Using this system, feeding performance and plant tissue damage by the root herbivore Bradysia (fungus gnat) were evaluated. Larval feeding was found to reduce Arabidopsis root biomass and water uptake.
Furthermore, we have characterized a root-specific terpene synthase AtTPS08, which is responsible for the constitutive formation of the novel volatile diterpene compound, rhizathalene, in Arabidopsis roots. Rhizathalene synthase is a class I diterpene synthase that has high affinity for the substrate geranylgeranyl diphosphate (GGPP) and is targeted to the root leucoplast. Expression of the β-glucuronidase (GUS) reporter gene fused to the upstream genomic region of AtTPS08 demonstrated constitutive promoter activity in the root vascular tissue and root tips. Using the established bioassay with Arabidopsis and Bradysia larvae, in aeroponic culture we could show that roots deficient in rhizathalene synthesis were more susceptible to herbivory. Our work provides in vivo-evidence that diterpene compounds are involved in belowground direct defense against root-feeding insects.
Future work is still required to improve our understanding of plant root defense. This study has provided a basis for future investigations on the biochemistry, molecular regulation and defensive function of Arabidopsis root chemicals in interaction with both above- and belowground herbivores (and pathogens).Ph. D.
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Höller, Ulrich. "Isolation, biological activity and secondary metabolite investigations of marine-derived fungi and selected host sponges." [S.l.] : [s.n.], 1999. http://www.gbv.de/dms/bs/toc/271061243.pdf.
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Jenkins, Kelly Matthew. "Chemical investigations of marine filamentous and zoosporic fungi and studies in marine microbial chemical ecology /." Diss., Connect to a 24 p. preview or request complete full text in PDF format. Access restricted to UC campuses, 1998. http://wwwlib.umi.com/cr/ucsd/fullcit?p9907830.
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Sarikaya, Bayram Özlem [Verfasser], Gerhard [Akademischer Betreuer] Braus, Stefanie [Akademischer Betreuer] Pöggeler, and Heike [Akademischer Betreuer] Krebber. "Role of methyltransferases in fungal development and secondary metabolite production / Özlem Sarikaya Bayram. Gutachter: Gerhard Braus ; Stefanie Pöggeler ; Heike Krebber. Betreuer: Gerhard Braus." Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2014. http://d-nb.info/1047706903/34.
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Libor, Benjamin [Verfasser]. "Implementation of the novel high-throughput Fungal Isolation one step Device method FIND and secondary metabolite extraction from the isolate Heydenia cf. alpina / Benjamin Libor." Bonn : Universitäts- und Landesbibliothek Bonn, 2021. http://d-nb.info/1239730047/34.
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West, Lauren Kelly. "An Approach to Enhance Secondary Metabolite Production of Endosymbiotic Fungi Through the Incorporation of Resin into Culture Media." Thesis, The University of Arizona, 2011. http://hdl.handle.net/10150/145097.
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Muszkieta, Laetitia. "Identification de nouveaux réseaux de régulation surexprimés dans l'appressorium du champignon phytopathogène Magnaporthe grisea." Thesis, Lyon 1, 2011. http://www.theses.fr/2011LYO10026.
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Magnaporthe grisea est responsable de la pyriculariose du riz, principale maladie de cette céréale. L’entrée du champignon dans la plante hôte se fait via l’appressorium. La différenciation de cette structure résulte d’une réorientation génétique et métabolique, et nécessite une régulation génétique fine. Une étude transcriptomique comparant les stades mycélium et appressorium a permis de montrer qu’environ 1300 ORFs sont surexprimées au stade appressorial. Ce transcriptome a permis l’identification de 32 gènes codant pour des facteurs de transcription pour lesquels dix mutants de délétion ont été générés et caractérisés. L’étude de leur pouvoir infectieux a révélé que le mutant délété du gène TF7 présente une pathogénie réduite de 70% sur plant d’orge. De plus, ce mutant est incapable de former des appressoria sur membrane artificielle sauf en présence d’un inducteur chimique (1,16- hexadecanediol). Par ailleurs, lorsque les appressoria sont formés, ils éclatent au bout de 14 heures. Cette altération peut être compensée par l’addition de sorbitol comme osmoprotecteur. Ce mutant est hypersensible à la Nikkomycine Z, un inhibiteur de la chitine synthase suggérant une altération du métabolisme pariétal. Un transcriptome différentiel réalisé à partir d’appressoria sauvages et mutés différenciés sur membrane de Téflon a révélé que des gènes impliqués dans le métabolisme de la chitine sont sous-exprimés dans le mutant ΔTf7. Le facteur de transcription Tf22 dont la délétion conduit à une réduction de 70% de la pathogénie sur riz a également fait l’objet d’une attention particulière. En effet, la recherche d’homologie a montré la présence de deux protéines homologues à Tf22 chez les deux champignons phytopathogènes S. nodorum et C. nicotianae et au-delà de la conservation d’un cluster potentiel de gènes du métabolisme secondaire, identifié chez C.nicotianae. La caractérisation du mutant a montré que l’expression de ce cluster potentiel est régulée négativement par le gène TF22Magnaporthe grisea is responsible for rice blast, the major disease of rice. The entry of the fungus in the host plant is via a specialized cell called appressorium. The differentiation of this structure results from a genetic and metabolic shift, and requires fine control mechanisms. A transcriptomic study comparing vegetative mycelium and appressorium mature stage, characteristic of the pre-penetration step was realized. In order to identify new regulatory networks specific of the appressorial differentiation, we focused on 32 genes encodi. Ten deletion mutants of transcription factor genes were generated and characterized. The study of their infectivity revealed that the TF7 gene deleted mutant has a reduced pathogenicity of 70% on barley plant resulting from an inability to penetrate the plant surface. Moreover, unlike the parental strain, this mutant is unable to form appressoria on artificial membrane except in the presence of a chemical inducer (1.16-hexadecanediol). Moreover, when appressoria are formed, they burst after 14 hours. This alteration can be compensated by a sorbitol solution acting as an osmoprotectant. This mutant is hypersensitive to nikkomycin Z, a chitin synthase inhibitor suggesting an alteration of parietal metabolism. A differential transcriptome was conducted comparing wild and mutated appressoria differentiated on Teflon membrane revealed that genes involved in chitin metabolism are dependent on the transcription factor Tf7. A second transcription factor Tf22 whose, deletion leads to a reduction of 70% of pathogenesis on rice, has also been studied. Indeed, the homology search showed the presence of two proteins homologous to Tf22 for S. nodorum and C. nicotianae. Beyond the conservation of the transcription factor, we observed the conservation of a potential cluster of genes of secondary metabolism identified in C.nicotianae. The characterization of the mutant revealed that expression of this potential cluster is negatively regulated by the gene TF22
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Nascimento, Isabela Maria do. "Estudo químico de duas linhagens de fungos endofíticos com atividade ao fitopatógeno Colletotrichum sp." Universidade de São Paulo, 2015. http://www.teses.usp.br/teses/disponiveis/11/11138/tde-29092015-111043/.
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Os fungos endofíticos constituem uma fonte promissora para descoberta de novos compostos ativos de interesse biotecnológico. Nas últimas décadas, os fungos endofíticos têm recebido atenção no meio científico devido aos compostos que produzem, com estruturas únicas e atividades biológicas de relevância em diversas áreas, inclusive agronômica. Um dos maiores problemas nesta área são as doenças causadas por fungos fitopatogênicos que acometem as culturas. Dentre elas, a antracnose é uma das principais doenças que atinge diversas culturas no Brasil, gerando grandes perdas agrícolas, principalmente as culturas do morango, guaraná e citrus, causadas por fungos do gênero Colletotrichum. Esse trabalho teve como objetivo investigar extratos de linhagens dos fungos endofíticos Fusarium e Penicillium, isolados de folhas do guaranazeiro, com atividade à fitopatógenos do gênero Colletotrichum isolados das culturas de guaraná, morango e citrus. Para tanto foi realizado uma triagem inicial de duas linhagens de fungos, dentre nove, por ensaios biológicos in vitro de cultivo pareado na atividade antifúngica aos três fitopatógenos. As duas linhagens selecionadas, apresentaram os maiores índices de antagonismo contra os três fitopatógenos e foram identificadas como Fusarium sp e Penicillum pinophilum. Os extratos e as frações semipurificadas se mostraram com grande potencial na inibição do crescimento do fitopatógeno do gênero Colletotrichum, causador da antracnose, que provoca grandes perdas nas culturas de importância econômica do Brasil.Endophitic fungi are a promising source for discovery of new active compounds of biotechnological interest. In recent decades, endophytic fungi have received attention in the scientific community because of compounds they produce, with unique structures and biological activities of relevance in several areas, including agronomy. One of the greatest problems in this is diseases caused by phytopathogenic fungi that affect the crops. Among them, anthracnose is a major disease that affects several crops in Brazil, generating large agricultural losses, especially strawberry crops, guarana and citrus caused by fungi of the genus Colletotrichum. This study investigated endophytic fungi strains of Fusarium and Penicillium extracts isolated from guarana leaves, with activity to phytopathogens of the genus Colletotrichum isolated from guarana crops, strawberry and citrus. For that, it was conducted an initial screening of two strains of fungi, among nine, using in vitro biological assays of cultivation paired in antifungal activity to the three pathogens. The two strains selected had the highest rates of antagonism against the three phytopathogens and were identified as Fusarium sp and Penicillium pinophilum. Extracts and semi-purified fractions were shown with great potential to inhibit pathogen growth of the genus Colletotrichum, which causes anthracnose and, thus, great losses to crops of economic importance in Brazil.
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Cheeseman, Kevin. "Aspects of Penicillium genomics : Molecular combing genome assembly, genetic exchange in food and potential for secondary metabolite production." Thesis, Paris 11, 2013. http://www.theses.fr/2013PA112280/document.
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Les Penicilliums sont des champignons filamenteux appartenant au genre Ascomycota. Ces champignons ont été utilisés par l’homme pour la production de nourriture depuis des siècles. Plus récemment, ils ont aussi été utilisés dans l’industrie biotechnologique pour la production de composés chimiques d’intérêts pharmaceutiques. Certaines espèces de Penicillium sont par ailleurs des moisissures contaminants certains aliments, d’autres sont des pathogènes de plantes, y compris de certains fruits. Leur génomique est globalement peut connue. Dans cette étude, nous avons analysé les génomes de deux espèces nouvellement séquencées, Penicillium roqueforti et Penicillium camemberti. Nous reportons ici le développement d’une nouvelle méthodologie pour l’amélioration et la validation d’assemblage de génomes en utilisant une technologie permettant l’observation de molécules d’ADN unique, le Peignage Moléculaire. En utilisant cette méthode, nous avons amélioré l’assemblage de Penicillium roqueforti. Ce manuscrit décrit aussi de multiples occurrences d’un transfert horizontal d’un ilot génomique de plus de cinq cent kilobases entre plusieurs Penicillium. Ce cas de transfert horizontal indique une fréquence d’échange latéral de matériel génétique plus forte qu’attendue. Enfin nous présentons un inventaire préliminaire du potentiel génomique pour la production de métabolites secondaires dans ces importants Penicillium alimentairesPenicillium are filamentous fungi belonging to the Ascomycota genus. Penicillium species have been used by Man for centuries in food making processes. More recently they have also been used in the biotechnology industry for the production of compounds of pharmaceutical interest. Some Penicillium species are food spoilage agents, pathogens of plants including fruits. Aspects of their genomics are largely unknown. In this study, we analysed the genomes of two newly sequenced species, Penicillium roqueforti and Penicillium camemberti. Here we report the development of a new methodology for improving and validating genome assembly using an original single DNA molecule technology, Molecular Combing. Using this methodology we were able to produce a high quality genome assembly of Penicillium roqueforti. This work also reports the multiple and recurrent horizontal transfer of a large genomic island of over half a megabase between several Penicillium species. This horizontal transfer indicates a higher frequency of lateral genetic exchange between cheesemaking fungi than previously expected. Finally, we present an early assessment of the genomic potential for secondary metabolite production in these important food associated penicilliums
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Sieber, Christian Martin Konrad [Verfasser], Hans-Werner [Akademischer Betreuer] Mewes, and Kirsten [Akademischer Betreuer] Jung. "Predicting virulence factors in filamentous fungi: Regulation and evolution of secondary metabolism gene clusters / Christian Martin Konrad Sieber. Gutachter: Hans-Werner Mewes ; Kirsten Jung. Betreuer: Hans-Werner Mewes." München : Universitätsbibliothek der TU München, 2015. http://d-nb.info/1067558845/34.
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Chagas, Fernanda Oliveira das. "Cultura mista, manipulação química e genética de micro-organismos: estratégias para a diversificação do metabolismo secundário." Universidade de São Paulo, 2014. http://www.teses.usp.br/teses/disponiveis/60/60138/tde-28102014-202425/.
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Recentes estudos genômicos têm mostrado que vários fungos e bactérias possuem um potencial biossintético superior à quantidade de me tabólitos secundários já isolados desses micro-organismos. A descoberta de produtos naturais inéditos e bioativos é limitada pela impossibilidade dos micro-organismos expressarem to das as suas rotas biossintéticas em laboratório. Assim, estratégias alternativas para i nduzir a produção de produtos naturais microbianos são necessárias. A utilização de cultur as mistas de micro-organismos é uma estratégia que vem sendo recentemente utilizada, na tentativa de mimetizar condições mais naturais de crescimento. Além disso, a adição de mo duladores químicos e epigenéticos às culturas microbianas também pode potencialmente est imular a produção de compostos de interesse, seja por ativar mecanismos celulares em resposta à condição de estresse, ou por alterar a taxa de transcrição de certos genes, em f unção de mudanças no grau de enovelamento da cromatina. Alternativamente, a indu ção de certos genes, e até mesmo a diversificação do metabolismo secundário, podem ser conseguidos através de engenharia genética, pela manipulação direta de genes de inter esse. A linhagem endofítica Alternaria tenuissima SS77, selecionada para os experimentos de modulaçã o química e epigenética, teve seu metabolismo secundário alterado após o tra tamento com diferentes moduladores. Provavelmente, o efeito observado ocorreu em função de uma eliciação inespecífica dos diferentes moduladores. Além disso, o cultivo misto desse fungo com o fungo endofítico Nigrospora sphaerica SS67 , isolada da mesma planta hospedeira ( Smallanthus sonchifolius ), levou ao isolamento de dois novos policetídeos, da classe das perilenequinonas, juntamente com um já relatado na literatura científica. Para r ealizar os cultivos microbianos mistos, envolvendo uma linhagem bacteriana e uma fúngica, t rês linhagens de actinobactérias e cinco de fungos, todos endofíticos da planta Lychnophora ericoides , foram selecionadas. Alterações no perfil metabólico da cultura mista de Phomopsis sp FLe6 com Streptomyces albospinus RLe7 foram as mais evidentes e por isso a maioria das investigações foram focadas nessa cultura mista. Várias condições de cu ltivo foram testadas e diferentes resultados foram obtidos. Em alguns casos, o desenv olvimento da linhagem fúngica foi inibido pela bacteriana, e em outros, foi observado o inverso. Da mesma forma, houve acentuada inibição da produção de alguns metabólito s secundários na presença da linhagem desafiadora, mas também foi verificada a eliciação de outros. Os extratos das culturas simples desses micro-organismos também apresentaram relativas alterações nos perfis metabólicos em função das condições de cultivo. Os metabólitos produzidos pelo fungo Phomopsis sp FLe6 e pela actinobactéria Streptomyces albospinus RLe7 foram isolados e caracterizados. Os resultados mostram que as intera ções entre os micro-organismos endofíticos são bastante complexas, estando sujeita s a ação de diversos fatores externos que muitas vezes não podem ser pré-determinados. Po r isso, estabelecer um cultivo misto adequado, do ponto de vista da eliciação da produçã o de metabólitos secundários, pode requerer uma série de tentativas. Ainda assim, os r esultados almejados podem ser conseguidos utilizando essa estratégia. Diferenteme nte das linhagens endofíticas, manipuladas quimicamente através de diferentes estr atégias, a linhagem sequenciada de Fusarium heterosporum ATCC 74349, foi manipulada geneticamente para a co nstrução de um gene biossintético híbrido pks-nrps , contendo a porção nrps do gene híbrido da equisetina e um pks críptico de Aspergillus fumigatus . Era esperado que a linhagem hibridizada fosse capaz de produzir o metabólito se cundário geneticamente planejado, entretanto, após seu cultivo, esse produto não foi detectado nos extratos, e as possíveis razões são discutidas. Ainda que os resultados espe rados não tenham sido obtidos, estudos que contribuam para a ampliação do entendimento das megassintases fúngicas são de extrema valia.Recently, genetic studies have shown that several b acteria and fungi hold a greater biosynthetic potential than the amount of secondary metabolites isolated from these microorganisms. The discovery of novel bioactive na tural products is limited by the inability of microorganisms to express all their biosynthetic pa thways in laboratory conditions. Therefore, alternative strategies to induce the production of microbial natural products are required. Mixed cultures of microorganisms are a strategy tha t has been used to mimic more natural conditions of growth. Furthermore, the addition of chemical and epigenetic modulators to the microbial cultures can also stimulate the productio n of compounds by activating cellular mechanisms in response to stress conditions or by c hanging the transcription rate of certain genes, due to changes in the chromatin folding. Alt ernatively, the induction of some genes, and even the diversification of secondary metabolis m, can be achieved by genetic engineering, by manipulating genes of interest. The endophytic strain Alternaria tenuissima SS77, which was selected for the experiments of che mical and epigenetic modulation, had changed its secondary metabolism after treatment wi th different modulators. Probably, the observed effect was due to a nonspecific elicitatio n of those modulators. Moreover, the mixed cultures of this fungus with the endophytic fungus Nigrospora sphaerica SS67, isolated from the same host plant ( Smallanthus sonchifolius ), led to the isolation of two new polyketides, belonging to perylene quinone class, along with ano ther one already reported in the scientific literature. Three strains of actinobacteria and fiv e fungi, all endophytes of Lychnophora ericoides , were selected to grow in microbial mixed cultures comprising one bacteria and one fungus. Changes in the metabolic profile of the mix ed culture of Phomopsis sp. FLe6 with Streptomyces albospinus RLe7 were the most obvious, and then further studi es were focused on this mixed culture. Many culture conditions were analyzed and different results were obtained. In some cases, the development of the fun gal strain was inhibited by bacteria, and in other cases was observed the opposite. Similarly , there was a remarkable inhibition of the production of certain secondary metabolites in the presence of the challenging strain, but the eliciting of others was also observed. The extracts of the single cultures of these microorganisms also showed changes in metabolic pro files due to culture conditions. The metabolites produced by the fungus Phomopsis sp. FLe6 and the actinobacteria S. albospinus RLe7 were isolated and characterized. The results show that interactions between endophytic microorganisms are quite complex and are influenced by various external factors that often can not be previously determined. Theref ore, establishing a suitable mixed culture to elicit the production of secondary metabolites m ay require some attempts. Still, the expected results can be achieved using this strateg y. Unlike the endophytic strains, that was chemically manipulated by different strategies, the sequenced strain Fusarium heterosporum ATCC 74349 was genetically manipulated to construct a hybrid PKS-NRPS biosynthetic gene containing the NRPS portion of the hybrid gene of e quisetin and a cryptic PKS gene of Aspergillus fumigatus . It was expected that hybridized strain could be a ble to produce the secondary metabolite genetically planned, however, after its cultivation, this product was not detected in any extracts, and some possible reasons are discussed. Although the expected results have not been obtained, studies that contri bute to increasing the understanding of fungal megasynthases are extremely valuable
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Santos, Suikinai Nobre. "Bioprospecção de biomoléculas isoladas de fungos endofíticos de Combretum leprosum do bioma Caatinga." Universidade de São Paulo, 2012. http://www.teses.usp.br/teses/disponiveis/11/11138/tde-07012013-083504/.
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Os micro-organismos que habitam o interior das plantas (endofíticos ou endófitos) tornaram-se foco de interesse por estarem envolvidos na produção de compostos químicos como enzimas, alcalóides, antibióticos, anticancerígenas e diferentes metabólitos. Os ecossistemas de regiões tropicais tem sido alvo de busca de compostos naturais por causa da riqueza de espécies e nichos ecológicos presentes nestas comunidades. O objetivo deste trabalho o isolamento, identificação e a bioprospecção de fungos endofíticos obtidos de Combretum leprosum e a detecção nos extratos de planta e micro-organismos da presença do composto combretastatin (CA4). Folhas, galhos, frutos e raízes de C. leprosum foram coletados de cinco estados dentro da zona de semiárido brasileiro: Bahia, Piauí, Ceará, Paraíba, Rio Grande do Norte. Partes das amostras foram triturados e submetidos à maceração primeiramente em diclorometano, seguidos de tetrahidrofurano e acetona de acordo com Pettit et al.(1987) para possível extração da CA4. Além disso, para avaliação in vitro da atividade citotóxica e antimicrobiana foram realizadas extrações em acetato de etila, clorofórmio e metanol. Foram detectados a possível presença da CA4 em todos os órgãos das plantas extraídos com tetrahidrofurano e as maiores concentrações foram observadas nas folhas. A atividade antitumoral dos extratos vegetais apresentaram as maiores inibições contra carcinoma (ovário IC50 10µg/mL-1, rim IC50 8,7µg/mL-1 e mama IC50 14,1µg/mL-1) e glioma.IC50 13,5µg/mL-1. A outra parte das amostras (folhas, caules e raízes) foram desinfetadas, fragmentadas e colocadas em meios de cultivo (Martin, BDA, Agar água) por 60 dias, 28°C. Foram isolados 405 fungos endofíticos e 159 apresentaram atividade contra fitopatogênicos, 72% para Rhizoctonia solani e 28% para Pythium aphanidermatum. As vinte e três linhagens que apresentaram as melhores atividades antifitopatogênicas foram submetidas a crescimento em Czapec em cultura estacionaria, por 30 dias, a 28°C, os respectivos metabólitos foram obtidos em múltiplo (3.0 e 11.0) e avaliados a atividade antimicrobiana contra bactérias patogênicas e fungos. Quatro linhagens foram selecionadas, identificadas pelo sequenciamento da região 18S, CFE177 como Fusarium oxysporum, CFE03 como Hypocrea koningii, linhagem CFE108 como Aspesgillus oryzae e CFE391 como Fusarium solani e avaliadas in vitro pelos testes biológicos: atividade antitumoral, antioxidante e antimicobactéria. Os compostos produzidos por A. oryzae CFE108 apresentaram potencial para bioprospecção, e de acordo com as atividade citotóxicas as maiores ações foram contra as linhagens linfoma histiocística (J744), mieloma murino (B16F10) e baixa citotóxidade para carcinoma de bexiga (ECV304) e leucemia eritroblástica humana (k562) na concentração de 1mg/mL-1. Foram isolados dois compostos: SS-XL-32-01 identificado como bis-(2-etilhexil) ftalato (DEHP) e SS-XL-20-1 identificado como fenol, 2.2 metilenobis[6-(1,1-dimetiletil)-4- etil], ambos com atividade anticâncer para células HeLa com percentual de ate 98% e 71%, de morte, respectivamente. Alem disso, a modificação através da reação de metilação do composto SS-XL-32-1 resultou na quebra do anel aromático, formação de 4 subprodutos e perda da atividade, sendo um indicativo do sitio ativo da molécula responsável pela atividade observada. Portanto, fungos endofíticos de 18 plantas do semiárido brasileiro podem ser considerados fonte de bioprospecção para novas moléculas bioativas com atividade antitumoral.The micro-organisms that reside in the aerial tissues and roots of plants (endophytic or endophyte) became the focus of interest for being involved in the chemical production such as enzymes, alkaloids, antibiotics, anticancer and different metabolites. The ecosystems of tropical region have been targeted search of natural compounds because of the richness of species and ecological niches present in these communities. The aim of this work was the isolation, identification and bioprospection for endophytic fungi from Combretum leprosum and detection in extracts of the plant and micro-organisms for the presence of the combretastatin (CA4). Leaves, stems, fruits and roots of C. leprosum were collected from five states within the semi-arid zone of Brazil: Bahia, Piaui, Ceara, Paraiba, Rio Grande do Norte. Part of the samples were crushed and subjected to maceration in dichloromethane, followed by tetrahydrofuran and acetone according to Pettit et al. (1987) for extracting the possible CA4. Moreover, for in vitro evaluation of the cytotoxic and antimicrobial activity extractions were carried out in ethyl acetate, chloroform and methanol. Were detected the possible presence of CA4 all plant organs extracted with tetrahydrofuran and the highest concentrations were observed on the leaves. The antitumor activity of plant extracts showed the highest inhibition against carcinoma (ovary IC50 10µg/mL-1, kidney IC50 8.7 µg/mL-1 and breast IC50 14.1 µg/mL-1) glioma IC50 and 13.5 mg-/mL-1. The other part of the samples (leaves, stems and roots) were disinfected, fragmented and placed in culture media (Martin, PDA, water agar) for 60 days, 28°C. 405 Endophytic fungi were isolated and 159 showed activity against phytopathogenic, 72% for Rhizoctonia solani and 28% for Pythium aphanidermatum. Twenty-three strains that showed good activities antiphytopathogenic, were grow on medium Czapec in static culture, for 30 days at 28°C, the respective metabolites were obtained in multiples pH (3.0 and 11.0) and evaluated the antimicrobial activity against pathogenic bacteria and fungi. Four strains were selected, identified by sequencing the 18S region, CFE177 as Fusarium oxysporum, CFE03 as Hypocrea koningii, strain CFE108 as Aspesgillus oryzae and CFE391 Fusarium solani, and evaluated by in vitro biological tests: antitumor, antioxidant and antimicobactérium activity. The compounds produced by A. oryzae CFE108 had biological potential and in accordance with the cytotoxic activity, showed the highest activities against lymphoma lines (J744), murine myeloma (B16F10) and low cytotoxicity for carcinoma of the bladder (ECV304) and leukemia erythroblastic human (K562) in 1mg/mL-1 concentration. Two compounds were isolated: SS-XL-32- 01 identified as bis-(2-ethylhexyl)phthalate (DEHP), and SS-XL-20-1 as phenol 2.2methylenobis [6-(1,1-dimethylethyl) - 4-ethyl], both with anticancer activity for HeLa cells with a percentage of up to 98% and 71%, of death, respectively. In addition, modified by methylation reaction of the compound SS-XL-32-1 resulted in the breaking of the aromatic ring and result in formation of four product and loss of activity being indicative of the active site of the molecule can be the aromatic ring. Therefore, endophytic fungi in semiarid Brazil plant can be considered a source of bioprospection for new bioactive molecules with anticancer activity.
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Gerke, Jennifer [Verfasser], Gerhard [Akademischer Betreuer] Braus, and Axel [Akademischer Betreuer] Zeeck. "Secondary metabolism and development in the filamentous fungus Aspergillus nidulans - Activation of silent gene clusters and characterization of the SAM synthetase SasA / Jennifer Gerke. Gutachter: Gerhard Braus ; Axel Zeeck. Betreuer: Gerhard Braus." Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2012. http://d-nb.info/1042846286/34.
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Thieme, Karl G. [Verfasser], Gerhard H. [Akademischer Betreuer] Braus, Ralf [Gutachter] Ficner, and Rolf [Gutachter] Daniel. "The Zinc cluster transcription factor ZtfA is an activator of asexual development and secondary metabolism and regulates the oxidative stress response in the filamentous fungus Aspergillus nidulans / Karl G. Thieme ; Gutachter: Ralf Ficner, Rolf Daniel ; Betreuer: Gerhard H. Braus." Göttingen : Niedersächsische Staats- und Universitätsbibliothek Göttingen, 2018. http://d-nb.info/1161183159/34.
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Navarri, Marion. "Métabolites secondaires de champignons de sédiments marins profonds : criblages génétique et fonctionnel et caractérisation structurale de molécules antimicrobiennes." Thesis, Brest, 2016. http://www.theses.fr/2016BRES0127/document.
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La propagation des micro-organismes résistants aux antibiotiques menace le système mondial de santé publique. Pour lutter contre ce phénomène, le renouvellement des molécules utilisées en antibiothérapie est devenu une priorité mondiale. Les antibiotiques étant principalement d’origine microbienne, l’étude des micro-organismes et de leurs métabolites s’est donc renforcée et s’oriente vers des écosystèmes peu explorés comme les biotopes marins.Nous avons exploré les activités antimicrobiennes d’une collection de 183 champignons isoles de sédiments marins profonds et collectés entre 4 et 1884 mètres sous le plancher océanique. Le potentiel de production de métabolites de cette collection a été révélé par un criblage génétique ciblant les PolyKetide synthase (PKS), les Non-Ribosomal Peptide Synthetase (NPRS), les TerPene Synthase (TPS) et les hybrides PKS-NRPS. Après avoir regroupé les isolats en fonction de leur profil MSP PCR, 110 ont été sélectionnés pour un criblage fonctionnel, montrant une forte proportion de champignons filamenteux antimicrobiens (32%).Après extraction et fractionnement, les composés bioactifs de 3 souches ont été caractérisés aux niveaux structural et fonctionnel. Ainsi, O. griseum UBOCC-A-114129 produit la fuscine, la dihydrofuscine, la secofuscine et la dihydrosecofuscine, P. bialowiezense UBOCC-A-114097 produit l’acide mycophénolique et Penicillium sp. produit UBOCC-A-114109 la rugulosine.Parallèlement, des analyses en LC-HRMS, réalisées sur des extraits fongiques, ont révélé un grand nombre de métabolites non décrits dans les bases de données. Les champignons des sédiments marins constituent donc un réservoir de structures originales à explorerThe spreading of antimicrobial resistant microorganisms jeopardizes global health caresystem. To counteract this threat the renewal of antibiotic molecules is a global priority. Antibioticcompounds are mainly originated from microorganisms, so microorganisms and their secondarymetabolites received an increasing interest. The search for new natural antimicrobial compoundsfrom microorganisms gained untapped ecosystems as marine biosphere.We investigated the antimicrobial properties of a fungal collection. The 183 fungal isolateswere collected from deep subseafloor sediment and isolated between 4 and 1,884 meters belowthe seafloor. Secondary metabolites production potential was studied for all isolates in thecollection by screening genes coding PolyKetide Synthase (PKS), Non-Ribosomal Peptide Synthetase(NRPS), TerPene Synthase (TPS) and hybrid PKS-NRPS. After isolates dereplication according to theirMSP-PCR fingerprinting, an antimicrobial screening was performed for 110 isolates, highlighting ahigh proportion of filamentous fungi with antimicrobial properties (32%).After extraction and bio-guided fractionation bioactive metabolites isolated from 3 strains,were characterized in a structural and functional manner: O. griseum UBOCC-A-114129 producedfuscin, dihydrofuscin, secofuscin and dihydrosecofuscine, P. bialowiezense UBOCC-A-114097synthetized mycophenolic acid and Penicillium sp. UBOCC-A-114109 produced rugulosin.In the meantime, LC-HRMS analysis, performed on fungal extracts, showed a great proportionof metabolites not detected in interrogated databases. So, deep subseafloor fungi, represent anuntapped reservoir of original structures to explore
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Blomberg, Patrik. "Non-target Effects of Genetically Modified Trees." Doctoral thesis, Umeå : Department of Ecology and Environmental Science, Umeå Universitet, 2007. http://urn.kb.se/resolve?urn=urn:nbn:se:umu:diva-1348.
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Kristina, Tešanović. "Биолошка активност и хемијски састав аутохтоних врста гљива Coprinus comatus (O.F. Müll.) Pers. Gray, 1797 и Coprinellus truncorum (Scop.) Redhead, Vilgalys & Monclavo, 2001." Phd thesis, Univerzitet u Novom Sadu, Prirodno-matematički fakultet u Novom Sadu, 2017. https://www.cris.uns.ac.rs/record.jsf?recordId=104928&source=NDLTD&language=en.
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У оквиру ове докторске дисертације испитана је биолошка активност екстраката плодних тела и потопљених култура (мицелије и филтрата) аутохтоних врста гљива Coprinus comatus и Coprinellus truncorum. Такође, испитан је метаболизам фосфата мицелија обе врсте употребом нуклеарно магнетне резонантне спректроскопије (31Р NMR), утицај ванадијума на метаболизам фосфата као и идентификација облика ванадата присутних у ћелији мицелије (51V NMR). Утврђена је антирадикалска и антиоксидативна активност етанолних,метанолних и водених екстраката гљива при чему су се екстракти потопљених култура издвојили по антирадикалској, а екстракти плодних тела по антиоксидативној активности. Екстракти потопљених култура истакли су се и у погледу антибактеријске активности, где се као најпотентнији показао хлороформски екстракт филтрата потопљене културе C. comatus. Такође, етанолни екстракт филтрата потопљене културе C. comatus показао се као најпотентнији у анти-ацетилхолинестеразној активности у односу на конвенционални лек донепезил. Испитан је и утицај екстраката на вијабилност ћелијских линија HepG2 (хумане хепатома ћелије) и Rin-5F (ß ћелије панкреаса пацова).Спектрофотометријским методама одређен је укупан садржај фенола и флавоноида у већини анализираних екстраката.LC/MS идентификацијом и квантификацијом фенолних киселина уочена је разлика између фенолних једињења присутних у плодном телу, мицелији и филтрату потопљене културе. Екстракти потопљених култура бележе већи број и већи садржај једињења. Укупан садржај протеина одређен само у воденим екстрактима, а укупан садржај угљених хидрата у полисахаридним екстрактима.Употребом Фуријеве инфрацрвене спектроскопске методе (FTIR) детектоване су везе између угљених хидрата присутних у полисахаридним екстрактима, а планарном хроматографијом показано је да екстракти плодног тела и филтрата врсте С. truncorum, као и екстракт плодног тела врсте C. comatus, садрже велику количину D-глукозе, док екстракт мицелије C. truncorum, баш као и екстракти филтрата и мицелије C. comatus, садрже највише галактозе. Квалитативном и квантитативном елементарном анализом (ААS) утврђен је виши садржај калијума и гвожђа у анализираним узорцима. GC-МS идентификацијом и квантификацијом масних киселина указано је на значајно присуство линолне киселине код обе врсте. Како за аутохтону врсту C.truncorum постоји мало података у литератури, подаци о њеном хемијском саставу могу се сматрати иновативним.Компаративним прегледом биолошке активности и хемијског састава екстраката плодног тела и мицелије и филтрата (потопљених култура) указано је да су анализирани екстракти извори биоактивних супстанци са медицинским потенцијалом, а потопљене културе датих гљива представљају атрактивне кандидате за даља биотехнолошка истраживања.U okviru ove doktorske disertacije ispitana je biološka aktivnost ekstrakata plodnih tela i potopljenih kultura (micelije i filtrata) autohtonih vrsta gljiva Coprinus comatus i Coprinellus truncorum. Takođe, ispitan je metabolizam fosfata micelija obe vrste upotrebom nuklearno magnetne rezonantne sprektroskopije (31R NMR), uticaj vanadijuma na metabolizam fosfata kao i identifikacija oblika vanadata prisutnih u ćeliji micelije (51V NMR). Utvrđena je antiradikalska i antioksidativna aktivnost etanolnih,metanolnih i vodenih ekstrakata gljiva pri čemu su se ekstrakti potopljenih kultura izdvojili po antiradikalskoj, a ekstrakti plodnih tela po antioksidativnoj aktivnosti. Ekstrakti potopljenih kultura istakli su se i u pogledu antibakterijske aktivnosti, gde se kao najpotentniji pokazao hloroformski ekstrakt filtrata potopljene kulture C. comatus. Takođe, etanolni ekstrakt filtrata potopljene kulture C. comatus pokazao se kao najpotentniji u anti-acetilholinesteraznoj aktivnosti u odnosu na konvencionalni lek donepezil. Ispitan je i uticaj ekstrakata na vijabilnost ćelijskih linija HepG2 (humane hepatoma ćelije) i Rin-5F (ß ćelije pankreasa pacova).Spektrofotometrijskim metodama određen je ukupan sadržaj fenola i flavonoida u većini analiziranih ekstrakata.LC/MS identifikacijom i kvantifikacijom fenolnih kiselina uočena je razlika između fenolnih jedinjenja prisutnih u plodnom telu, miceliji i filtratu potopljene kulture. Ekstrakti potopljenih kultura beleže veći broj i veći sadržaj jedinjenja. Ukupan sadržaj proteina određen samo u vodenim ekstraktima, a ukupan sadržaj ugljenih hidrata u polisaharidnim ekstraktima.Upotrebom Furijeve infracrvene spektroskopske metode (FTIR) detektovane su veze između ugljenih hidrata prisutnih u polisaharidnim ekstraktima, a planarnom hromatografijom pokazano je da ekstrakti plodnog tela i filtrata vrste S. truncorum, kao i ekstrakt plodnog tela vrste C. comatus, sadrže veliku količinu D-glukoze, dok ekstrakt micelije C. truncorum, baš kao i ekstrakti filtrata i micelije C. comatus, sadrže najviše galaktoze. Kvalitativnom i kvantitativnom elementarnom analizom (AAS) utvrđen je viši sadržaj kalijuma i gvožđa u analiziranim uzorcima. GC-MS identifikacijom i kvantifikacijom masnih kiselina ukazano je na značajno prisustvo linolne kiseline kod obe vrste. Kako za autohtonu vrstu C.truncorum postoji malo podataka u literaturi, podaci o njenom hemijskom sastavu mogu se smatrati inovativnim.Komparativnim pregledom biološke aktivnosti i hemijskog sastava ekstrakata plodnog tela i micelije i filtrata (potopljenih kultura) ukazano je da su analizirani ekstrakti izvori bioaktivnih supstanci sa medicinskim potencijalom, a potopljene kulture datih gljiva predstavljaju atraktivne kandidate za dalja biotehnološka istraživanja.
The biological activity of extracts of basidiocarps (fruiting bodies) and submerged cultures (mycelium and filtrate) of autochthonous mushroom species Coprinus comatus and Coprinellus truncorum was examined. Furthermore, the metabolism of phosphate of mycelia of both types was studied using nuclear magnetic resonance spectros-copy ( 31 R NMR), the influence of vanadium on phosphate metabolism and the identification of vanadate oxidation states present in the mycelia cell ( 51 V NMR). The antiradical and antioxidant activity of methanolic, ethanolic and water fungal extracts was determined. Extracts of submerged cultures achieved the best anti- radical activity while fruit body extracts showed the best antioxidant activity. Extracts of submerged cultures also highlighted in terms of antibacterial activity, where the chloroform extract of the submerged culture C. comatus showed as the most potent. Also, the ethanolic extract of the submerged culture of C. comatus was found to be most relevant in anti-acetylcholinesterase activity compared with the conventional donepezil drug. The influence of extracts on the viability of cell lines HepG2 (human hepatocytes cells) and Rin-5F (ß pancreatic cells of the rat) was also examined.Spectrophotometric methods determined the total con-tent of phenol and flavonoids in most of the analyzed extracts.The LC/MS identification and quantification of phenolic acids revealed the difference between the phenolic compounds present in the fruiting body, mycelium, and the submerged culture filtrate. Extracts of submerged cultures record a greater number and higher content of compounds.The total content of proteins determined only in water extracts and the total content of carbohydrates in poly-saccharide extracts. Using the Fourier infrared spectro-scopic method (FTIR), the links between the sugar pre-sent in the polysaccharide extracts were detected, and planar chromatography showed that the extracts of the fruiting body and the filtrate of type C. truncorum, as well as the extract of the fruiting body of the species C. comatus, contain a large amount of D-glucose, while the extract of the C. truncorum mycelia and mycelia of C. comatus, contain the most galactose. GC-MS identification and quantification of fatty acids indicated a significant presence of linoleic acid in both species, while qualitative and quantitative elemental analysis (AAS) has determined a higher content of potas-sium and iron in the analyzed samples. Since there is no data in the literature for the autochtho-nous species C. truncorum, the studies on its chemical composition can be considered advanced аs innovative. A comparative review of the biological activity and the chemical composition of the extracts of the fruiting body and mycelia and filtrates of medium of submerged cultures indicated that the extracts were analyzed by sources of bioactive substances with medical potential, and the submerged cultures of these mushrooms are attractive candidates for biotechnological research.
В рамках данной работы была исследованна биологическая активность экстракта плодородных тел и погружонных видов култур (мицелии и филтрата) автотоных видов грибов Coprinus comatus и Coprinellus truncorum. Также, исследованн метаболизм фосфата обеих видов мицелий с помощью ядерного магнитного резонанса спектроскопии (31Р ЯМР), влияние на содержание ванадия в метаболизме фосфата, а также идентификация формы ванадата присущего в клеток мицеллий (51V ЯМР). Установленная антирадикальная и антиоксидантная активность метанольных, этанольных и водных экстрактов гриб, причём выделяются экстракты погружённых культур по антирадикальной активности и экстракты плодородных тел по антиоксидантной активности.Экстракты погружённых культур выделялись и в плане антибактериальной активности, причем, наиболее мощным из филтратов показался экстракт хлороформа погруженной культуры C. comatus. А также этанольный экстракт филтрата погружённой культуры C. comatus оказался найболее мощным в анти-ацетихолинестеразной активностипо сравнению с традиционным лекарством донепезилом. Было исследовано и влияние экстрактов на виябильность клеток линий HepG2 (гуманые хепатома клетки) и Rin-5F (ß клетки поджелудочной железы крыс).Методом спектрофотометрии определена совокупность фенола и флавоноида в большинстве проанализированных экстрактах.С помощью ЛС ̸МС идентификации и квантификации фенолных кислот была замечена разница между соединениями фенола, присущих в плодородном теле, и мицелии, и филтрата погружённой культуры. Экстракты погружённых культур отражают больше количество и более высокое содержание соединений.Общее содержание белков выделен только в водяных экстрактах, и общее содержание углеводов в полисахаридных экстрактах. Используя инфракрасный метод спектроскопии Фурия (ИКМСФ) были обнаружены связи между сахарами, присущими в полисахаридных экстрактах, а планарной хромотографиой было показано, что экстракты плодородного тела и филтратов вида С. truncorum, а также и экстракты плодородного тела вида C. comatus содержат большое количество D-глюкозы, в то время как экстракт мицелии C. truncorum, именно как и экстракт фильтрата и мицелии C. comatus, содержат больше всего галактозы.GC-МS идентификацией и квантификацией жирных кислот показано значительное наличие линолевой кислоты у обоих видах. А качественным и квантитативным элементарным анализом установленно большее содержание калиума и железа в анализированых шаблонах.Из-за того, что для автохтонного вида C. truncorum практически не было данных в литературе, данные о её химическом составе можно считать прогрессивным и инновационным.Сравнительный анализ биологической активности и химического состава экстрактов плодородного тела и мицелии и фильтрат (погружённых культур) показаывает, что проанализированные экстракты — источники биологически активных веществ с медицинским потенциалом, и погружённые культуры данных гриб являются привлекательными кандидатами для биотехнологических исследований.
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Alves, Paula Cristina. "Triggering secondary metabolite biosynthesis: exploring the effects of ionic liquids in fungal metabolism." Doctoral thesis, 2016. http://hdl.handle.net/10362/43835.
Full textAbstract:
Filamentous fungi are able to synthesise an array of small molecules (secondary
metabolites), which are usually not essential for fungal growth but confer
competitiveness. As a consequence, numerous secondary metabolites remain cryptic
at the artificial conditions of cultivation in a research laboratory. Even in
Aspergillus nidulans, one of the most well studied fungi, numerous metabolites
remain unseen. Several strategies have been used to solve this knowledge gap, some
of which require prior knowledge of genomic sequences, relying on manipulation of
targeted genes encoding components of either secondary metabolism or regulatory
pathways. Other approaches may be applied also in less well characterised strains,
such as cultivation with other species/organisms or modification of the growth
media composition. (...)
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Caballero, Ortiz Silvia. "Fungal Responses to Grazers." Doctoral thesis, 2014. http://hdl.handle.net/11858/00-1735-0000-0023-98FF-8.
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Sarikaya, Bayram Özlem. "Role of methyltransferases in fungal development and secondary metabolite production." Doctoral thesis, 2014. http://hdl.handle.net/11858/00-1735-0000-0022-5E29-6.
Full textAbstract:
Pilzentwicklung und Sekundärmetabolismus werden durch Einwirkung von
Umwelteinflüssen von Regulatorproteinen kontrolliert. Das VeA Protein repräsentiert die
velvet-Domänen-Familie der Pilzregulatoren. VeA passt die sexuelle Entwicklung und den
dazu gehörenden Sekundärmetabolismus von Aspergillus nidulans an die Lichtverhältnisse
an. VeA bindet im Dunkeln an VelB und bildet schließlich den trimeren VelB-VeA-LaeA
(velvet) Komplex. VeA dient als Brückenprotein für das velvet-Domänen-Protein VelB als
Regulator der Entwicklung und die Methyltransferase LaeA als Regulator des
Sekundärmetabolismus. VelB kann mit VosA einen zweiten licht-regulierten Komplex
bilden, der die asexuelle Entwicklung reprimiert. Auch VosA gehört zur Familie der Velvet-
Proteine. LaeA kontrolliert die Bildung der VelB-VosA und VelB-VeA-LaeA Komplexe
während der Entwicklung. laeA Nullmutationen können nicht mehr auf Licht reagieren, was
ihre Schlüsselrolle als Regulatoren der Entwicklung unterstreicht. Die Abwesenheit von LaeA
führt zur Bildung von wesentlich kleineren Fruchtkörpern. Grund hierfür ist das Fehlen
runder Hülle-Zellen, die den jungen Fruchtkörper ernähren und in seiner Entwicklung
unterstützen. LaeA spielt damit eine dynamische Rolle während der morphologischen und
biochemischen Entwicklung des Pilzes, indem die Expression, Interaktion und die
Modifikation der velvet Regulatoren kontrolliert werden. Im zweiten Teil der Arbeit wurde
die VeA-Plattform für Protein-Protein Interaktionen weiter untersucht. VeA interagierende
Proteine (Vips) identifiziert in einen „Yeast-two-hybrid“ System führten zu einem trimeren
Methyltransferase-Komplex, der Signaltransduktion mit epigenetischer Kontrolle verbindet.
Der neuartige Komplex enthält das Plasmamembran-assoziierte Trimer VapA-VipC-VapB.
Das Dimer VipC-VapB ist über das FYVE-ähnliche Zinkfinger Protein VapA an die
Plasmamembran gebunden und ermöglicht dem nuklearen VelB-VeA-LaeA Komplex die
Aktivierung der Transkription der sexuellen Entwicklung. Sobald die Abkopplung vom VapA
stattgefunden hat, wird VipC-VapB zum Kern transportiert. VipC-VapB interagiert
physikalisch mit VeA, vermindert dessen Transport zum Kern und die Stabilität. Folglich
wird der Anteil des VelB-VeA-LaeA Komplexes im Kern reduziert. Die nukleare VapB
Methyltransferase vermindert die Entstehung des fakultativen Chromatins indem es die
Histon 3 Lysin 9 Methylierung (H3K9 me3) vermindert. Dies begünstigt die Aktivierung der
frühen Regulatorgene flbA und flbC, die dann das asexuelle Programm im Licht vorantreiben.
Der VapA-VipC-VapB Methyltransferase-Weg vereinigt die Kontrolle des Kernimportes und
der Stabilität von Transkriptionsfaktoren mit der Modifikation von Histonen. Erst dieses
komplexe Zusammenspiel unterschiedlicher Mechanismen erlaubt eine angemessene Antwort
für die Differenzierung des Pilzes.
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Kralj, Ana [Verfasser]. "Isolation of secondary fungal metabolites and their influence on sphingolipid metabolism / vorgelegt von Ana Kralj." 2007. http://d-nb.info/986814954/34.
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Meister, Cindy. "Interplay of the COP9 signalosome deneddylase and the UspA deubiquitinase to coordinate fungal development and secondary metabolism." Doctoral thesis, 2018. http://hdl.handle.net/21.11130/00-1735-0000-0003-C10F-3.
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Thieme, Sabine. "Insertion of an intrinsically disordered domain in VelB supports selective heterodimer formation of fungal velvet domain regulatory proteins in Aspergillus nidulans." Doctoral thesis, 2018. http://hdl.handle.net/11858/00-1735-0000-002E-E5FE-3.
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Dreyer, Jacqueline Paola Ilka [Verfasser]. "Global regulators of fungal secondary metabolism: molecular genetic characterization of Velvet in the β-lactam [beta-lactam] producer Acremonium chrysogenum / submitted by Jacqueline Paola Ilka Dreyer." 2007. http://d-nb.info/985241926/34.
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Lin, Cheng-Pan, and 林振邦. "Study on the Mechanisms of Anticancer Activity of a Fungal Secondary Metabolite, Cephalochromin, in Nasopharyngeal Carcinoma Cells." Thesis, 2009. http://ndltd.ncl.edu.tw/handle/80668015121706916992.
Full textAbstract:
碩士中國醫藥大學
藥學系碩士班
97
According to the recently information release from the Department of Health, the malignant tumor is the number 1 cause of death in Taiwan for the last 27 years. Although, several great improvement have been established for treating cancer, developing the drug resistance by the cancer cells causes the curative effect generally not good. Therefore, the new antitumor medicine''s research and development appear urgent. At present the source of antitumor agents is major isolated or derivatives from the plant or the animal, but recently, several studies demonstrated that the fungus also obtain the effective anti-tumor active compounds. In this study, we identified a fungal secondary metabolite, cephalochromin, which posseses a strong antitumoral activity against several human cancer cell lines. Since cephalochromin exerts comparably great anti-proliferative effect toward human nasopharyngeal carcinoma HONE-1 and NPC-TW01 cells, we therefore investigated the mechanisms of action of anticancer efficacy of cephalochromin in this type of cancer. Results demonstrated that cephalochromin induces cells arrest in the G1 phase in the time- and doseage-dependent manner. Significant appearance of sub-G1 population and Annexin V-positive cells indicates that cephalochromin-induced cell death proceeded through an apoptotic pathway. Furthermore, we found that cephalochromin only activate the caspase-8, but not caspase-9. The finding that cephalochromin-induced apoptosis through a memebrane-mediated mechanism was supported by up-regulated expression of Fas and FasL. Furthermore, up-regulation of p53 was found after cephalochromin exposure. Above results indicated that cephalochromin is a effective anticancer agent which could induced cancer cell apoptosis through a p53-mediated caspase-8 / Fas-FasL-dependent pathway and worth for further development.
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Tugizimana, Fidele. "Metabolite profiling of defence-related secondary metabolites in tobacco cells, in response to ergosterol, a steroid from fungal membranes." Thesis, 2012. http://hdl.handle.net/10210/8088.
Full textAbstract:
M.Sc.Plants have the ability to continuously respond to various stimuli which alter their physiology, morphology and development. These stimuli may be abiotic or biotic and range from essential to toxic in their effects. One of these stimuli is a steroid from fungal membranes, ergosterol (C28H44O), which does not occur in plants. Ergosterol acts as a pathogen-associated molecular pattern molecule and triggers defence mechanisms in plants, characterised by highly regulated and interrelated events that include the elicitation of the oxidative burst and expression of a number of defencerelated genes. However, the ergosterol-induced global cellular reprogramming of the host has not been fully investigated in all aspects. No metabolomic study has previously been conducted to elucidate, for instance, the effect of ergosterol on plant metabolism. A clear and broader understanding of the molecular mechanisms involved in plant : ergosterol interactions is of paramount importance, for it would open up possibilities of developing novel, more effective and sustainable strategies to control or eradicate fungal diseases in plants. In plants, the metabolome is a compilation of all primary and secondary metabolites. The latter are the final recipients of genetic information, and their levels can influence gene expression and protein stability. Metabolite patterns reveal the actual cellular dynamic environment. Hence, qualitative and quantitative measurements of extra- and intracellular metabolites yield insights into the cellular processes that control the biochemical phenotype of the cell, tissue or whole organism. Metabolomics, the most recent of the ‘omics’ approaches, is the holistic analysis of metabolites present within a biological system under specific physiological conditions. In the present study a metabolomic approach was used to elucidate and analyse changes in the metabolism of tobacco (Nicotiana tabacum) cells following ergosterol treatment. Special attention is given to sesquiterpenoids since the antimicrobial compounds (phytoalexins) isolated from plants within the Solanaceae are mostly bicyclic sesquiterpenoids. Suspension of tobacco cells were treated with different concentrations (0 - 1000 nM) of ergosterol and incubated for different time periods (0 - 24 h). A viability assay, based on the ability of viable cells to reduce 2,3,5- triphenyltetrazolium chloride (TTC), was used to determine whether cell death occurred due to ergosterol treatment. No loss of cell viability was observed over the concentration range and time periods used in this study, indicating that the observed responses were due to the treatment alone and possible secondary responses due to cell death could be excluded. Intracellular metabolites were extracted with two methods: a selective dispersive liquid-liquid micro extraction and a general methanol extraction. Chromatographic techniques (TLC/HPTLC, GC-FID, GC-MS, GC×GC-TOF-MS, UPLC-MS) and 1H NMR spectroscopy were used for quantitative and qualitative analyses. Multivariate data analyses (PCA and OPLS-DA models) were used to extract interpretable information from the multidimensional data generated from the aforementioned techniques.
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"Strain degeneration in Aspergillus parasiticus: A model system for variation in secondary metabolite producing filamentous fungi." Tulane University, 1991.
Find full textAbstract:
Strain variation ('degeneration') in fungi is characterized by changes in phenotype, attenuated virulence, reduced sporulation, and/or loss of secondary metabolite production. In this study, wild type and mutant strains of Aspergillus parasiticus (designated sec+) making aflatoxin and/or pigmented pathway intermediates were subjected to a protocol of serial transfer of mycelial macerates in a defined medium. Variant forms (designated sec$-$) exhibiting altered morphology, scanty sporulation, and lack of detectable amounts of secondary metabolite production were isolated after 5-12 transfers of mycelia. In contrast, when spores rather than mycelia were serially transferred, no $sec-$ forms were generated. The $sec-$ forms were stable and did not revert to sec+ phenotype upon subculturing on complete medium for 10 weeks. The DNA methylation patterns and, with one exception, the dry weights of the sec+ and $sec-$ strains were similar To conduct parasexual cycle analysis, heterokaryons between sec+ and $sec-$ forms with contrasting auxotrophic and spore color markers were produced. Spore color, auxotrophic markers, and the sec+ or $sec-$ phenotype failed to segregate in the heterokaryon test, suggesting that degeneration was not under cytoplasmic control. Qualitative and quantitative assays of heterozygous diploids showed similar aflatoxin production (155-157 $\mu$g/100 ml defined medium) in sec+/sec+ and sec+/sec$-$ diploids, indicating dominance of the sec+ phenotype Haploids were isolated from diploids using benomyl as the haploidization agent. The appearance of a few revertants of the $sec-$ forms from the sec+/sec$-$ crosses indicated that the aflatoxin pathway genes were present but turned off in these variants. To investigate the nature of non-expression, biotransformation experiments were conducted, where both sec+ and sec$-$ forms were fed with 200 $\mu$g of sterigmatocystin, an aflatoxin precursor. All the sec+ forms successfully biotransformed sterigmatocystin to aflatoxin. All the sec$-$ forms did not convert the precursor to detectable amounts of aflatoxin. This phenomenon of non-expression of aflatoxin pathway genes in the experimentally induced sec$-$ forms suggests that this fungus could be developed as a model system to understand the poorly understood process of strain degeneration in filamentous fungiacase@tulane.edu
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Höller, Ulrich [Verfasser]. "Isolation, biological activity and secondary metabolite investigations of marine-derived fungi and selected host sponges / von Ulrich Höller." 1999. http://d-nb.info/956675638/34.
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Gerke, Jennifer. "Secondary metabolism and development in the filamentous fungus Aspergillus nidulans - Activation of silent gene clusters and characterization of the SAM synthetase SasA." Doctoral thesis, 2012. http://hdl.handle.net/11858/00-1735-0000-000D-EF8A-9.
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Dirnberger, Benedict. "Proteomics of Aspergillus nidulans sexually differentiated cells." Doctoral thesis, 2018. http://hdl.handle.net/11858/00-1735-0000-002E-E4B4-D.
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Thieme, Karl G. "The Zinc cluster transcription factor ZtfA is an activator of asexual development and secondary metabolism and regulates the oxidative stress response in the filamentous fungus Aspergillus nidulans." Doctoral thesis, 2017. http://hdl.handle.net/11858/00-1735-0000-002E-E418-F.
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Crossman, Julia Stephanie. "Biomimetic apporaches to the synthesis of polyketide derived marine natural products (-)-Maurenone and the spiculoic acids /." 2007. http://catalogue.flinders.edu.au/local/adt/public/adt-SFU20080212.134949/index.html.
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Nahlik, Krystyna. "The COP9 signalosome of Aspergillus nidulans." Doctoral thesis, 2007. http://hdl.handle.net/11858/00-1735-0000-0006-B4EA-B.
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Bayram, Özgür. "Blue light-dependent development of the filamentous fungus Aspergillus nidulans." Doctoral thesis, 2007. http://hdl.handle.net/11858/00-1735-0000-0006-ACE3-5.
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