Characterizing Douglas-fir Tissue Colonization by the 'Sudden Oak Death' Pathogen, Phytophthora ramorum.

Characterizing Douglas-fir Tissue Colonization by the 'Sudden Oak Death' Pathogen, Phytophthora ramorum.
By Kathleen McKeever, M.S. and Dr. Gary Chastagner; Department of Plant Pathology, Washington State University Puyallup Research and Extension Center.

In 2001, Koch's Postulates confirmed Douglas-fir as a host for P. ramorum. Naturally-infected saplings have been observed in California forests, and studies on artificially inoculated Douglas-fir stems and shoots have established susceptibility parameters. Although naturally occurring stem infections in the U.S. have been limited to smaller diameter seedlings and saplings, P. ramorum stem cankers were recently observed on 8-year-old plantation grown Douglas-fir in Great Britain in 2009. All previous research has served to substantiate the importance of performing studies to characterize the behavior of P. ramorum in Douglas-fir and assess the potential risk that this pathogen poses to Douglas-fir ecosystems.

Artificial inoculation experiments were carried out to provide a thorough analysis of the colonization of Douglas-fir by P. ramorum. Goals of this research included determining which tissues are colonized by the pathogen, whether woody tissues are able to support sporulation, the likelihood of stem infections occurring on Douglas-fir with intact bark, and the viability of the pathogen in foliage. Methods employed included isolation, ELISA, and histological examination of stem tissues; qPCR and isolation to determine colonization efficacy and viability of the pathogen in needle tissues; and baiting studies to determine the ability of Douglas-fir bark to inhibit colonization of Rhododendron leaves by the pathogen.

ELISA results showed that proteins of the pathogen were detectable in the phloem, cambium, and superficial xylem, with infrequent detection in asymptomatic tissues. The pathogen was able to be isolated from all symptomatic woody tissues tested, but not from non-discolored tissues. ELISA and isolation techniques produced results that were highly positively correlated (r2=0.62, p=0.78), and histological observations paralleled information derived from these techniques. Douglas-fir bark reduced infection on Rhododendron leaf baits by up to 83 percent in the presence of inoculum. Results from foliage inoculations indicated that pathogen DNA is detectable using qPCR methods, but there was an inability to isolate the pathogen from needle tissue. Evidence of spore formation in woody stem tissues has not been observed.

It was concluded that the pathogen may be able to infect into the shallow xylem tissues of Douglas-fir in the presence of wounding, but there was no evidence of sporulation in tissues. The inability to isolate the pathogen from non-discolored tissues suggests that the proteins detected by ELISA outside of the lesion may be elicitins that are secreted in advance of hyphal colonization. The inability to isolate the pathogen from colonized needles may indicate the presence of chemical inhibitors that render the pathogen non-viable subsequent to initial infection. Finally, the ability of Douglas-fir bark to suppress infection of Rhododendron leaf baits suggests that the bark is inhibitory to P. ramorum and that infection of woody stem tissues through intact bark may be limited. Further research is required to identify the nature of this inhibition.

Characterization of P. ramorum colonization of Douglas-fir tissues is relevant to our current understanding of the behavior of the pathogen in different hosts and may enhance our ability to assess risk and create adequate regulations to thwart the spread of this exotic pathogen.

from the December COMTF newsletter