Thesis (Ph. D.)--University of Wisconsin--Madison, 1949. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 105-106).

Thesis (Ph. D.)--University of Wisconsin--Madison, 1943. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaf 27).

Fungi contribute to key processes in the sustainable function of terrestrial ecosystems including nutrient cycling and transport of water to plants. However, some fungal species are of interest because their infection of a susceptible host crop results in diseases negatively affecting food supply and quality. These diseases are expected to be influenced by rotation crops which could impact the development of plant pathogens and their genetic biodiversity. The objectives of this study were to assess the biodiversity of fungal pathogens in durum wheat, to identify the naturally occurring fungi that could have biocontrol potential, and to define the impact of crop rotation with non-cereal crops on fungal populations in durum wheat. For this purpose, in 2004, 2005, and 2006, soil and durum wheat plant samples were collected after preceding crops of <i>Pisum sativum</i> L. (pea), <i>Lens culinaris</i> Medik (lentil), <i>Cicer arietinum</i> L. (chickpea), <i>Brassica napus</i> L. (canola) and <i>Triticum turgidum</i> L. (durum) in a long term experimental site in Swift Current, Sakatchewan. Samples were analyzed using a combination of traditional cultivation techniques and polymerase chain reaction (PCR), sequencing, and denaturing gradient gel electrophoresis (DGGE) techniques. <p>Fusarium species, known as the causal agent of Fusarium head blight (FHB) and Fusarium damaged kernels (FDK) were among the most ubiquitous and abundant in durum tissues. The most prevalent of all Fusarium at the study site were <i>F. avenaceum, F. reticulatum</i>, and <i>F. tricinctum</i>. Other recovered potential fungal pathogens belonged to the genera Bipolaris, Phaeosphaeria, Pyrenophora, Cladosporium, Epicoccum, Alternaria, Cladosporium, Arthrinium, Nigrospora, and Microdochium. Principal component analysis revealed negative correlations between Acremonium, Chaetomium, Penicillium, and pathogenic Fusarium, Bipolaris, Pyrenophora, and Alternaria. These isolates could be antagonistic, and their potential as biocontrol agents against pathogens colonizing durum wheat in the semiarid Saskatchewan should be assessed.<p>Crop rotation had a limited impact on the abundance of fungal pathogens. Fusarium torulosum was less abundant in durum following canola while Bipolaris sorokiniana was less abundant in durum following pea. Even if no single crop rotation reduced significantly the prevalence of F. avenaceum in durum wheat, results suggest that a successful control of this important pathogen requires an integrated approach using diversified rotations.

Fungi contribute to key processes in the sustainable function of terrestrial ecosystems including nutrient cycling and transport of water to plants. However, some fungal species are of interest because their infection of a susceptible host crop results in diseases negatively affecting food supply and quality. These diseases are expected to be influenced by rotation crops which could impact the development of plant pathogens and their genetic biodiversity. The objectives of this study were to assess the biodiversity of fungal pathogens in durum wheat, to identify the naturally occurring fungi that could have biocontrol potential, and to define the impact of crop rotation with non-cereal crops on fungal populations in durum wheat. For this purpose, in 2004, 2005, and 2006, soil and durum wheat plant samples were collected after preceding crops of <i>Pisum sativum</i> L. (pea), <i>Lens culinaris</i> Medik (lentil), <i>Cicer arietinum</i> L. (chickpea), <i>Brassica napus</i> L. (canola) and <i>Triticum turgidum</i> L. (durum) in a long term experimental site in Swift Current, Sakatchewan. Samples were analyzed using a combination of traditional cultivation techniques and polymerase chain reaction (PCR), sequencing, and denaturing gradient gel electrophoresis (DGGE) techniques. <p>Fusarium species, known as the causal agent of Fusarium head blight (FHB) and Fusarium damaged kernels (FDK) were among the most ubiquitous and abundant in durum tissues. The most prevalent of all Fusarium at the study site were <i>F. avenaceum, F. reticulatum</i>, and <i>F. tricinctum</i>. Other recovered potential fungal pathogens belonged to the genera Bipolaris, Phaeosphaeria, Pyrenophora, Cladosporium, Epicoccum, Alternaria, Cladosporium, Arthrinium, Nigrospora, and Microdochium. Principal component analysis revealed negative correlations between Acremonium, Chaetomium, Penicillium, and pathogenic Fusarium, Bipolaris, Pyrenophora, and Alternaria. These isolates could be antagonistic, and their potential as biocontrol agents against pathogens colonizing durum wheat in the semiarid Saskatchewan should be assessed.<p>Crop rotation had a limited impact on the abundance of fungal pathogens. Fusarium torulosum was less abundant in durum following canola while Bipolaris sorokiniana was less abundant in durum following pea. Even if no single crop rotation reduced significantly the prevalence of F. avenaceum in durum wheat, results suggest that a successful control of this important pathogen requires an integrated approach using diversified rotations.

Title page, table of contents and abstract only. The complete thesis in print form is available from the University of Adelaide Library. / Barley net-and spot-form of net blotch, caused by two formae of the hemibiotrophic fungus Pyrenophora teres, are two of the major diseases affecting barley crops worldwide. In this study, the method of suppression subtractive hybridisation was used to isolate barley epidennal genes that were differentially expressed in the early stages of both net blotch incompatible compared to compatible interactions. As a result, two subtracted libraries of cDNA clones comprising mainly of gene transcripts of low abundance were generated. Quantitative real-time PCR was employed to verify and profile the differential expression of forty-five subtracted transcripts during the first 48 hours of infection, resulting in the identification of twenty-eight clones that were pathogen-induced and differentially expressed. These clones were grouped into one of eight clusters depending on the kinetics of their expression, and they included groups of genes that were up-regulated early (within 3 hai) and later (24 hai) in both barley-P. teres incompatible interactions. Among the differentially expressed clones were those with sequence homology to genes that encode proteins involved in calcium signal perception (e.g. a calcineurin B-like protein), detoxification (e.g. multidrug transporters), carbohydrate metabolism (e.g. an invertase), and signal transduction (e.g. protein kinases). Furthennore, the expression profiles generated for each individual gene cluster were similar for both net-and spot-form interactions, indicating that the resistance-associated defence response against both pathogens may be mediated by the same molecular mechanism. The differentially expressed genes are discussed with respect to their potential functional role in contributing to net blotch disease resistance. In addition, a model detailing early events that may take place in the barley-P. teres incompatible interaction is presented. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1286782 / Thesis (Ph.D.) -- University of Adelaide, School of Agriculture, Food and Wine, 2007

No description available.

Graduation date: 1998

Pyrenophora teres f. teres is the causal agent of net form net blotch of barley. P. teres f. teres is prevalent globally across all barley growing regions and globally is the most devastating foliar disease of barley. Though economically important, the molecular mechanism whereby P. teres f. teres causes disease is poorly understood and investigations into these mechanisms have been hindered by a lack of genomic resources. To set a genomic foundation for P. teres f. teres the reference isolate 0-1 was sequenced and assembled using PacBio single molecule real-time (SMRT) sequencing and scaffolded into 12 chromosomes to provide the first finished genome of P. teres f. teres. High confidence gene models were generated for the reference genome of isolate 0-1 using a combination of pure culture and in planta RNA sequencing. An additional four P. teres f. teres isolates were sequenced and assembled to the same quality as the reference isolate 0-1 and used in a comparative genomic study. Comparisons of the five P. teres f. teres isolates showed a two-speed genome architecture with the genome being partitioned into core and accessory genomic compartments. Accessory genomic compartments clustered in sub-telomeric regions of the P. teres f. teres genome with a majority of previously identified quantitative trait loci (QTL) associated with avirulence/virulence being spanned by these accessory regions. Using these genomic resources, with a bi-parental mapping population and a natural population for QTL analysis and genome wide association study (GWAS), respectively, we identified a candidate gene for the previously mapped AvrHar. QTL analysis identified a locus extending off the end of P. teres f. teres chromosome 5 and GWAS analysis identified significant associations with a gene encoding a small secreted protein. The candidate AvrHar gene was validated using CRISPR-Cas9-RNP gene disruption in parental isolates 15A and 0-1. Disruption of AvrHar in isolate 15A did not result in a phenotypic change while disruption of the 0-1 allele resulted in a complete loss of pathogenicity. This is the first identification of an effector from P. teres f. teres validated using CRISPR-Cas9-RNP gene editing. / North Dakota Barley Council

Pyrenophora teres, the causal agent of net blotch of barley (Hordeum vulgare L.), exists in two forms; P. teres f. teres and P. teres f. maculata. Both forms induce a combination of brown necrotic spots and extensive chlorosis in susceptible barley cultivars. Although a number of low molecular weight compounds (LMWCs) have been previously isolated from P. teres culture filtrates, they only induced certain components of symptoms. Fungal metabolites were extracted from culture filtrates of both forms of the pathogen and separated into low (<3kDa) and high molecular weight compounds (HMWCs, >10 kDa) with each fraction inducing a component of the net blotch symptoms in a barley leaf toxicity assay. Inactivation of both LMWCs (<1kDa) and HMWCs resulted in loss of activity confirming their potential role in symptom development. Low molecular weight compounds induced chlorosis and water soaking but not the brown necrotic spots or lesions usually seen during the infection of barley by P. teres. The high molecular weight compounds (>10 kDa) induced the brown necrotic spots or lesions with no chlorosis evident. Further characterisation of LMWCs showed that they are not host specific while HMWCs exhibited host specificity. LMWCs were purified and further analysed using high voltage paper electrophoresis, staining and mass spectrometry. Electrophoretic properties and staining of the LMWCs with ninhydrin indicated that both forms of P. teres produced similar LMWCs in the conditions grown. Each form produced eight ninhydrin-positive compounds with the samerelative mobilities. Each individual compound was shown to induce chlorosis in excised barley leaves. All compounds except the one isolated in this study appear to be derivatives of or are the previously described compounds; N-(2-amino-2carboxyethyl) aspartic acid (Toxin A), aspergillomarasmine A, anhydroaspergillomarasmine A and aspergillomarasmine B. The exception is a bioactive UV absorbing LMWC which appears to be a reductive conjugation of the α-keto acid of phenylalanine with Toxin A. The HMWCs (>10kDa) were proteinaceous since they were identifiable using Coomassie staining. Additionally, the loss of activity that occurred with incubation at 40, 60, and 80 °C for 30 and 60 min followed a pattern fairly typical for protein denaturation. Further, treatment with protease decreased their phytotoxicity in proportion to the amount of enzyme used. Enzyme and heat treatment of proteins extracted from each form showed that proteins of P. teres f. teres are more resistant to heat and enzyme treatment compared with those of P. teres f. maculata. This suggests the protein(s) involved in symptom induction by P. teres f. teres and P. teres f. maculata are different which contributes to the difference in the symptom expression during the interaction between the pathogens and barley. Proteinaceous metabolites extracted from P. teres f. teres and P. teres f. maculata ranged from 10 to 100 kDa. Fractions purified using gel filtration had biological activity when they contained eight proteins when extracted from P. teres f. maculata (90, 80, 75, 55, 48, 35, 14 and 12 kDa) and six proteins when extracted from P. teres f. teres (90, 80, 55, 48, 14 and 12 kDa). Additionally, intercellular washing fluids (IWF) extracted from barley plants inoculated with both forms of P. teres, contained proteins of the same size as those in the biologically active fractions extracted from culture filtrates of P. teres f. maculata (80, 14 and 12 kDa) and P. teres f. teres (80, 48 and 14 kDa). Automated MS/MS sequencing of the biologically active proteins showed no resemblance to the sequences or conserved domain information available in public databases and as a consequence, these proteins were considered as novel proteins for P. teres. However, exact short matches with fragments resulting from the 80, 48 and 14 kDa proteins, showed considerable homology with ATP-binding cassette (ABC) transporters and their components, cellulases, serine proteinases as well as some hypothetical proteins isolated from different fungal species. Reaction of six plant species including one susceptible barley cultivar (Sloop) and one resistant line (CI9214) to P. teres showed that partially purified proteins induce the symptoms selectively in barley cultivars where the proteinaceous metabolites only induced brown necrotic spot/lesions in barley with a greater response seen on the susceptible cultivar Sloop when compared to the resistant line CI9214. No symptoms were seen on other plant species employed in this study suggesting that the proteinaceous metabolites isolated in this study are host specific phytotoxins. This research has allowed the first isolation of proteinaceous host-specific toxins from P. teres as well as the identification of a UV-sensitive LMWC phytotoxin not previously described. Proteinaceous toxins induced brown necrotic spots/lesions specific to the host while the LMWCs induced chlorosis in a number of different plant species. This contributes significantly to the body of knowledge defining how symptoms are caused during the pathogenicity process in the interaction between P. teres and barley. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1297672 / Thesis (Ph.D.) -- University of Adelaide, School of Agriculture, Food and Wine, 2007

Tan spot of wheat is an economically significant disease caused by the fungal pathogen, Pyrenophora tritici-repentis. Certain races of the fungus secrete Ptr ToxA (ToxA), a 13.2 kDa proteinaceous host-selective toxin that is responsible and sufficient to cause disease in susceptible wheat varieties. Disease symptoms develop only when the ToxA gene in the fungus and a single gene in the wheat host are expressed. The understanding of this gene-for-gene interaction could be instrumental towards control of the disease and is also being developed as a model system for understanding host-pathogen interactions. Here, this effort is given a solid structural foundation through crystallographic analysis of the ToxA structure. The ToxA structure was solved at 1.65 Å resolution using the anomalous signal from inherently present sulfur atoms. The monomeric toxin adopts a β-sandwich fold of two anti-parallel β-sheets composed of four strands each. The mapping of existing mutation data onto the structure reveals that a sequence of Arg- Gly-Asp(RGD) and surrounding residues required for activity are present on a solvent-exposed loop thereby making them potential candidates for recognition events that are required for ToxA activity. Unexpectedly, after a simple circular permutation, the ToxA structure is topologically identical to the classic mammalian RGD containing fibronectin type III (FnIII) domain, and furthermore the RGD residues are topologically equivalent. These results support the hypothesis that ToxA, like FnIII, interacts with an integrin-like receptor on the host plant cell surface. There has been a renewed interest in the method of using the anomalous signal from sulfur atoms to solve protein structures. As a spin-off of the structure solution work, the data were systematically analyzed to study the effects of crystal decay, resolution and data redundancy on the ability to locate the sulfur positions and subsequent phasing of the protein. The analyses show that the choices made about data redundancy and resolution limits may be crucial for the structure determination and that anomalous correlation coefficients are helpful indicators in making these choices. / Graduation date: 2006