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United States Department of Agriculture

Agricultural Research Service

Research Project: ENHANCING PATHOGEN DETECTION AND CROP PROTECTION IN SUGARBEET USING MOLECULAR TECHNOLOGIES
2009 Annual Report


1a.Objectives (from AD-416)
Three of the most significant field diseases of sugarbeet in the U.S. are root rot, caused by Aphanomyces cochlioides; Rhizomania, caused by a fungal/viral complex; and wilt, caused by the sugarbeet cyst nematode, Heterodera schachtii, and concomitant infection by Fusarium fungi. The objectives of this project are to investigate methods to reproduce the field diseases of these pathogens in controlled environments, to develop qualitative and quantitative detection reagents and protocols for these organisms, and to determine genetic changes in viruses of the Rhizomania complex that condition heightened virulence to sugarbeet. Since the incorporation of natural genetic resistance into crops remains the most cost-effective strategy for disease control, an additional objective of the project is to obtain molecular genetic tags for disease resistance genes in sugarbeet in collaborative studies with ARS sugarbeet geneticists and pathologists.


1b.Approach (from AD-416)
Gradients of saturation across seedbeds will be tested as a means to evaluate sugarbeet varieties with known resistance to Aphanomyces cochlioides using stand loss as a measure of disease severity. Protocols for the inoculation of sugarbeet with Polymyxa betae will be modified to select for clonal isolates of the organism, an aspect lacking in past studies on this pathogen. Probe primers will be designed to perform in conjunction with specific primer sets in the development of real-time PCR (qPCR) methods for quantifying these pathogens in soil and plant samples. Disruptions (insertions) in the chromosomes of beet black scorch virus and beet necrotic yellow vein virus will be engineered in efforts to determine the role of virus genes in pathogen virulence. Plants typed for either resistance or susceptibility to the sugarbeet cyst nematode and Fusarium stalk blight will be subjected to DNA fingerprinting for generation of molecular markers linked to resistance genes.


3.Progress Report
Research to design primers to distinguish Aphanomyces cochlioides and Rhizoctonia solani has resulted in the successful development of species and sub-species specific primers. Using these primers, PCR conditions have been optimized for specificity and reliability to distinguish R. solani sub-species AG 2-2 IIIB and AG 2-2 IV, and A. cochlioides from other sugarbeet pathogens. Methods to isolate and purify apoplastic fluid from Cercospora beticola-infected sugarbeet leaves have been optimized. In collaborative research with colleagues at North Dakota State University, a multi-year study to determine the occurrence and frequency of mating type genes in field isolates of C. beticola has been completed. Research to determine the environmental parameters affecting the severity of R. solani infection has been completed in collaboration with colleagues at North Dakota State University.


4.Accomplishments
1. Defining environmental factors controlling sugarbeet Rhizoctonia root rot infection to optimize fungicide timing and efficacy. Rhizoctonia root and crown rot is an increasing problem in sugarbeet growing areas in the United States. Chemical control is the primary means to control disease caused by this fungus but is much more effective if applied prior to infection. In collaboration with North Dakota State University, research conducted by the Sugarbeet & Potato Research Unit, Northern Crop Science Laboratory, Fargo, ND has defined the environmental parameters (soil moisture and soil temperature) that are required for sugarbeet root rot infection. This information will allow growers to better estimate the likelihood of Rhizoctonia infection and initiate chemical control measures for optimal disease control thereby improving sugarbeet yield and producer profitability. In addition, the identification of environmental parameters optimal for R. solani infection will facilitate testing of new and existing fungicides for improved disease control.

2. Tracking fungicide resistance in the sugarbeet leaf spot pathogen. Cercospora beticola is the pathogen that causes the economically devastating sugarbeet leaf spot disease. Field isolates of C. beticola exhibit high levels of variability in appearance and, more importantly, fungicide resistance. Studies in Europe have shown that C. beticola populations have a high degree of genetic variation and an equal distribution of mating type genes which suggests that sexual reproduction may contribute to the appearance and spread of fungicide resistance. However, no sexual stage of this fungus has been found. In collaboration with plant pathologists at North Dakota State University, research over the past three years from the Sugarbeet & Potato Research Unit, Northern Crop Science Laboratory, Fargo, ND has identified the mating types of field isolates of Cercospora found in the north-central U.S. and found that, while sexual reproduction is likely, there exists no correlation between mating type and fungicide resistance. These results suggest that fungicide resistance occurs naturally in Cercospora and resistance management strategies are necessary to minimize the spread of fungicide resistance in this economically important sugarbeet pathogen. These results may help explain why fungicide resistance in C. beticola populations is an endemic threat to sugarbeet growers in this country. This study represents the first report of the occurrence and distribution of C. beticola mating type genes in North America.


6.Technology Transfer

Number of the New/Active MTAs (providing only)1

Review Publications
Bolton, M.D. 2009. Primary Metabolism and Plant Defense - Fuel for the Fire. Molecular Plant-Microbe Interactions. 22(5):487-497.

van Esse, H.P., van't Klooster, J.W., Bolton, M.D., Yadeta, K.A., Van Baarlen, P., Boeren, S., Vervoort, J., de Wit, P.J.G.M., Thomma, B.P.H.J. 2008. The Cladosporium fulvum Virulence Protein Avr2 Inhibits Host Proteases Required for Basal Defense. The Plant Cell. 20:1948-1963.

deSilva, A.P., Bolton, M.D., Nelson, B.D. 2009. Transformation of Sclerotinia Sclerotiorum with the Green Fluorescent Protein Gene and Fluorescence of Hyphae in Four Inoculated Hosts. Plant Pathology. 48:487-496.

DeTemmerman, N., Anfinrud, M., Meulemans, M., Rick, K., Burkholz, A., DeBruyne, E., Weyens, G., Barnes, S., Horemans, S., Lefebvre, M., Bolton, M.D. 2009. Rhizomania Resistance in the Tandem Sugar Beet Variety. International Sugar Journal. 111(1325):313-317.

Last Modified: 11/27/2014
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