Rapid New Diagnostic LAMP (Loop-Mediated Isothermal Amplification) Assays to Distinguish Among the Four Lineages of Phytophthora ramorum.

Sudden oak death (SOD) is caused by Phytophthora ramorum, an invasive oomycete pathogen. This pathogen is of major regulatory concern for nurseries, horticulture, and forestry in the United States and around the world. Three of the 12 identified lineages of P. ramorum currently occur in the United States (NA1, NA2, and EU1) affecting wildland forests and nurseries. Rapid identification and lineage determination is essential to accelerate management decisions, detect introductions of new lineages, and control the spread of SOD. The objective of this study was to develop and validate diagnostic tools to rapidly identify P. ramorum and distinguish among the four common lineages of the pathogen and to accelarate management decision making. The loop-mediated isothermal amplification (LAMP) assays developed here are species specific with no cross reaction to common Phytophthora species found in Oregon, California, and Washington. The lineage-specific assays unambiguously distinguish among the four common clonal lineages. These assays are sensitive and able to detect P. ramorum DNA ranging in concentration from 30 to 0.03 ng/μl depending on the assay. These assays work effectively on a variety of sample types including plant tissue, cultures, and DNA. They have been integrated into the SOD diagnostic process in the forest pathology lab at Oregon State University. To date, 190 samples have been correctly identified from over 200 field samples tested for lineage determination. The development of these assays will help managers in forestry and horticulture identify and rapidly respond to new outbreaks of P. ramorum.