Graduate Thesis Or Dissertation

Growth properties of induced mutants of the common wheat-bunt fungus, Tilletia caries, in axenic culture and in planta

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  • Genetic studies of Tilletia spp. have been hampered by the lack of genetic markers. The work discussed here is a first report of the isolation of genetically marked strains of J. caries and their use in wheat plant infection studies. Procedures for the isolation of auxotrophic and drug-resistant mutants in haploid strains of J. caries were developed. Preliminary determination of the nuclear condition of secondary sporidia showed that spores were typically mononucleate. Additionally, characterization of the kinetics of secondary sporidial formation in liquid culture provided essential information concerning the extent of the logrithmic phase of growth and spore generation rates. Log-phase sporidia of two monokaryotic, compatible strains of race T-l were mutagenized with nitrosoguanidine. Two "non-leaky" nutritional mutants were isolated and characterized; one required adenine, the other uracil. Five "leaky" nutritional mutants were also isolated. One was identified as a glycine auxotroph, and the other four had mutations in pathways for general nitrogen metabolism. Additionally, secondary sporidia of T. caries were tested for their growth response to twelve different drugs in attempts to isolate drug-resistant strains. Only mutants resistant to cycloheximide were detected. Pairings of compatible auxotrophic strains produced mycelial growth on a minimal medium whereas unpaired strains typically failed to grow. Growth in the absence of nutritional supplements was presumed to be due either to 'forced' heterokaryon formation or syntrophism (i.e., crossfeeding). Inability to successfully isolate viable hyphal tips from colonies arising from paired auxotrophic strains precluded proof of heterokaryosis. Hypodermic inoculations with compatible auxotrophic, drug-resistant, and wild type strains failed to result in teliospore production in wheat plants of the susceptible spring wheat var. 'Red Bobs'. J. caries was successfully reisolated as four morphologically distinct colony or sector types from 38 plants that failed to head normally. It was also isolated from a small number of mature plants showing no signs of disease. All auxotrophic and drug-resistant strains were successfully recovered as individual haploid isolates from outgrown colonies on wheat plant tissue. Inability to success fully isolate viable hyphal tips precluded proof of heterokaryosis. Consequently, the nuclear condition, i.e., heterokaryon or homokaryon, of the outgrown fungus was not determined.
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