01860nas a2200157 4500008004100000245013300041210006900174300001400243490000700257520128500264100002501549700001701574700001601591700002301607856007201630 2004 eng d00aDetection and quantification of Phytophthora ramorum from California forests using a real-time polymerase chain reaction assay-00 aDetection and quantification of Phytophthora ramorum from Califo a1075-10830 v943 a

The timely and accurate detection of pathogens is a critical aid in the study of the epidemiology and biology of plant diseases. In the case of regulated organisms, the availability of a sensitive and reliable assay is essential when trying to achieve early detection of the pathogen. We developed and tested a real-time, nested polymerase chain reaction (PCR) assay for the detection of Phytophthora ramorum, causal agent of sudden oak death. This technique then was implemented as part of a widespread environmental screen throughout California. The method here described is sensitive, detecting less than 12 fg of pathogen DNA, and is specific for P. ramorum when tested across 21 Phytophthora spp. Hundreds of symptomatic samples from 33 sites in 14 California counties were assayed, resulting in the discovery of 10 new host species and 23 infested areas, including 4 new counties. With the exception of a single host, PCR-based discovery of new hosts and infested areas always was confirmed by traditional pathogen isolations and inoculation studies. Nonetheless, molecular diagnostics were key in early pathogen detection, and steered the direction of further research on this newly discovered and generalist Phytophthora species.

1 aHayden, Katherine, J1 aRizzo, David1 aTse, Justin1 aGarbelotto, Matteo uhttp://apsjournals.apsnet.org/doi/abs/10.1094/PHYTO.2004.94.10.1075