Kati Orr University of Idaho, Moscow, Idaho M.S. Soil Microbiology Plant, Soil, and Entomological Sciences Department Advisor: Guy R. Knudsen Graduated: December 2003 Work Phone: (208) 885-5769 Email: orr9345@uidaho.edu

M.S. Thesis Title:

Image analysis of GFP-tagged Trichoderma harzianum in soil and a molecular comparison of conserved genes in T. harzianum and Fusarium oxysporum.

M.S. Thesis Topic:

Proliferation of Trichoderma harzianum determined by traditional plating techniques and image analysis
Many fungi exist in soil as hyphae, conidia and chlamydospores; therefore, changes in soil populations are often difficult to interpret with traditional culturing methods.  Hyphal growth of Trichoderma harzianum occurs during favorable environmental conditions while sporulation increases when conditions become unfavorable or stressful, such as restrictive matric potentials or reduced nutrient availability.  The dilution plating technique is the most widely used and accepted method in soil microbiology for enumerating soil microorganisms.  However, in the case of T. harzianum, direct cell counts can be significantly larger than viable counts due to the presence of numerous resting spores, generating inaccurate estimates of population growth.   Microscopic analysis, on the other hand, can differentiate among the different fungal propagules and provide a closer estimation of actively growing biomass than can dilution plating.  T. harzianum (ThzID1-M3), with a gene encoding for green fluorescent protein (GFP), was illuminated under UV light and measured with imaging software to determine biovolume using hyphal-lengths and diameters.  Biomass can then be estimated using appropriate density conversion factors.

ThzID1-M3 expressing GFP

ThzID1-M3 in soil

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