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United States Department of Agriculture

Agricultural Research Service

Title: Examining Interactions Between Legumes and Aphamonyces euteiches with Real-time PCR

Authors
item Vandemark, George
item Ariss, Jennifer

Submitted to: Australasian Plant Pathlogy Society
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: November 6, 2006
Publication Date: March 6, 2007
Citation: Vandemark, G.J. and J.J. Ariss. 2007. Examining interactions between legumes and aphanomyies enteiches with real-time PCR.Australasian Plant Pathology 36:102-108.

Interpretive Summary: The majority of plant diseases are caused by fungi, and previous methods for quantifying fungi in plant tissue, such as staining of fungal biomass with dyes or detection of fungal-specific proteins, are very limited in precision. At the USDA-ARS, Prosser, WA, a new method for quantifying the amount of pathogen present in infected plants has been developed based on a “real-time fluorescent PCR” assays. These assays can be used to both detect and quantify the soilborne plant pathogens Aphanomyces euteiches and Phytophthora medicaginis, which are responsible for root rot disease of alfalfa and other legumes. The assays are very precise and for four different isolates of A. euteiches, a very significant positive correlation has been observed between the amount of pathogen DNA detected with these assays and the severity of disease in infected alfalfa. Similarly, for three of five isolates of A. euteiches, significant positive correlation were observed between the amount of pathogen DNA detected in infected roots and disease severity in infected peas. With a single isolate of P. medicaginis, signficant correlations were also observed betwen pathogen DNA accumulation in infecetd roots and disease severity in infected plants. Experiments were also performed to examine the accumulation of A. euteiches in resistant and susceptible pea populations at several different time points (7, 10, and 14 days) during the progress of disease. At all times, significantly more A. euteiches DNA was detected in the susceptible population than in the resistant population. In both resistant and susceptible populations the quantity of A. euteiches DNA reached a maximum at 7 days, and then decreased in a fairly linear manner over time. These observations provide evidence that A. euteiches is a biotrophic organism that primarily produces mycelia early in the disease cycle. These results suggest that control measures be targeted to disrupt the pathogen early in the process of disease to reduce the production of oospores that serve as subsequent field inoculum. The results also suggest that it may be possible to select resistant plants as early as 7 days after infection, indicating that this technology can be used to rapidly accelerate the development of new pea varieties with desired disease resistance profiles.

Technical Abstract: Real-time PCR has many applications in the study of host-pathogen interactions including improving the resolution of disease screening programs in selecting highly resistant plants. Identifying the earliest time in the disease process that real-time PCR can be used to reliably identify resistant genotypes will optimize applications of these assays in resistance breeding. Experiments were performed to examine the accumulation of A. euteiches in resistant and susceptible pea populations at several different time points (7, 10, and 14 days) during the progress of disease. At all times, significantly more A. euteiches DNA was detected in the susceptible cultivar Bolero than in the resistant germplasm 90-2079. The earliest significant correlation between disease severity and pathogen DNA quantity was observed 14 days after inoculation. In both resistant and susceptible populations the quantity of A. euteiches DNA reached a maximum at 7 days, and then decreased in a fairly linear manner over time. These observations provide evidence that A. euteiches is a biotrophic organism that primarily produces mycelia early in the disease cycle. These results suggest that control measures be targeted to disrupt the pathogen early in the process of disease to reduce the production of oospores that serve as subsequent field inoculum.

Last Modified: 12/20/2014
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