Project Number: 5302-13000-011-17
Project Type: Reimbursable

Start Date: Oct 01, 2012
End Date: Sep 30, 2014

Objective:
1) Identify pathogenic Pythium species using RFLP and sequencing based assay. 2) Provide a faster means of estimating pathogenic Pythium species populations using real time PCR. 3) Develop internal control to determine if PCR inhibitors influences the amplification of pathogenic Pythium species target sequences.

Approach:
Plant and soil samples will be collected randomly from under the diseased calla lily plants from the Golden State Bulb Growers in Moss Landing, CA. Infected roots will be cut in 5-mm segment and placed on Pythium selective media plates and will be established in Potato Dextrose Agar after 2 days. Pathogenicity test will be performed in the greenhouse. Identification of Pythium species will be done using PCR-RFLP of the mitochondrially encoded cytochrome oxidase II (cox II) gene followed by sequencing of cox II gene. Soil DNA extraction will be performed using procedures identified previously. Quantification of pathogenic Pythium species will be accomplished using real time PCR quantification assay since it offers a rapid, sensitive, and specific method for the diagnosis of plant pathogens in soil samples. Inoculum densities will also be determined using soil plating. Evaluation of DNA extraction procedures will be done with different soil types.