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United States Department of Agriculture

Agricultural Research Service

Research Project: Development and Application of An Epidemiological Framework for Management of Phytophthora Ramorum in Nurseries

Location: Horticultural Crops Research

Project Number: 2072-22000-039-27
Project Type: Interagency Reimbursable Agreement

Start Date: Oct 01, 2013
End Date: Sep 30, 2018

Objective:
Our approach addresses the epidemiology of soilborne sources of P. ramorum by assessing two aspects of the disease cycle crucial for disease establishment: sporulation by pathogen sources in the soil, and subsequent infection of hosts at the soil surface. Immediate objectives to be accomplished at the NORSDUC facility for the current fiscal year include: 1. Determine the relative importance and durability of two forms of inoculum, infested leaf debris or ‘naked’ chlamydospores, in initiating infection of rhododendron roots or leaves at the soil-plant interface. 2. Assess survival and sporulation potential of inoculum buried within soils over time.

Approach:
Objective 1. Inoculum will be introduced into soils as either inoculated rhododendron leaf disks, or ‘naked’ chlamydospores at the start of the experiment. To prevent drying, all inoculum will be covered in 1 cm of soil and will be routinely watered. Periodically over time these sources will be tested for their ability to initiate infection of plant tissues. We will assess infection of both leaves and roots. For each assessment period, uninfected rhododendron leaf disks will be placed atop infested soil for 3 days and watered sufficiently to maintain leaf wetness. The disks will be collected, incubated for 7 days, then surface sterilized and plated in selective media to assess for colonization by P. ramorum. To test the ability of inoculum within soil to infect root balls, during the same 3 day period as the leaf assays we will place potted rhododendron plants on the soil surface. After the exposure period the root balls will be gently removed from the pot and washed. Wounded rhododendron leaves will then be placed at the bottom of the pot, and the plants will be returned and generously watered. After 4 days, the bait leaves will be retrieved and assessed for colonization by P. ramorum as described above. Objective 2. Infected rhododendron leaf disks will be placed in small, nylon mesh bags. These will either be left on the soil surface, or buried in soil columns at depths ranging between 1 cm and 10 cm. At each sampling period, a subsample of leaf disks will be retrieved and assessed for the potential to produce inoculum: disks will be rinsed, then placed in Falcon tubes with deionized water to stimulate the formation of sporangia, which will be filtered and quantified by using microscopy and qPCR. After testing for inoculum-production, all samples will be surface sterilized and plated in selective media to assess survivability of P. ramorum in leaf debris at each depth.

Last Modified: 11/23/2014
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