2012 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.
This is the final report for Project 5442-22000-042-00D, which has been replaced by bridging Project 5442-22000-047-00D. Substantial results were realized over the course of this project. The ability to reliably reproduce sugarbeet diseases in controlled settings is foundational for studying pathogen biology and disease resistance screening. Environmental parameters were assessed to optimize isolation and inoculation procedures for four important sugarbeet pathogens. Molecular techniques were developed to detect pathogens in diseased tissue, which helps to streamline the ability to quantify disease. To gain insight into the molecular interaction between sugarbeet and the important pathogens Cercospora beticola and Aphanomyces cochlioides, proteomic techniques were used isolate secreted proteins from both pathogens. Such proteins may be useful for future disease resistance screening strategies. Studies on the population biology of C. beticola showed that the fungus has many hallmarks of sexual reproduction and populations are genetically diverse, which may explain why the pathogen is able to readily develop fungicide resistance. In collaboration with ARS scientists in Fargo, ND, treatments with the plant hormone jasmonic acid were identified that reduced root rot to three important pathogens of sugarbeet. To find genes that are critical for this resistance response, next generation sequencing was utilized to identify sugarbeet genes differentially expressed after exposure to jasmonic acid, which may be valuable biomarkers for resistance screening. Research to determine the environmental parameters minimally and optimally required for infection with the soilborne pathogen Rhizoctonia solani were completed. This information is useful for sugarbeet growers, as it helps to determine the optimal timing for fungicide applications. The concepts and discoveries generated from this project plan have led to a more thorough understanding of sugarbeet pathology and host-pathogen interactions. The overall impact of this project is that sugarbeet growers and industry have new information on which to guide decisions concerning sugarbeet disease management, which will help to increase yield and profits.
Identification of the sugarbeet disease organism in North Dakota. The sugarbeet cyst nematode is a devastating disease of sugarbeet. Knowledge of the occurrence and distribution of any plant pathogen is necessary for disease control and prevention. In collaboration with plant pathologists at North Dakota State University, ARS researchers at Fargo, ND were the first to identify fields containing the sugarbeet cyst nematode in North Dakota. Since the sugarbeet cyst nematode is also an important pathogen of canola, this information is important for sugarbeet and canola growers so they can minimize future impacts and reduce inadvertent spread of this pathogen.
Identification of a genetic mutation associated with fungicide resistance. Leaf spot, caused by the fungus Cercospora beticola, is the most important foliar disease of sugarbeet and is controlled in part by the application of fungicides. However, fungicide resistance in C. beticola is a constant threat to sugarbeet growers and results in financial losses due to application of fungicides with reduced efficacy. In collaboration with plant pathologists at North Dakota State University, ARS scientists in Fargo, ND were the first to identify strains of C. beticola with resistance to a common class of fungicides. In addition, a specific genetic mutation that is associated with strobilurin resistance was identified. Knowledge of fungicide resistance will help guide decisions on choosing the optimum fungicide for effective disease control, which will decrease production costs and increase yields.
Bolton, M.D., Secor, G.A., Rivera, V., Weiland, J.J., Rudolph, K., Birla, K., Rengifo, J., Campbell, L.G. 2012. Evaluation of the potential for sexual reproduction in field populations of Cercospora beticola from USA. Fungal Biology. 116:511-521.
Zhong, S., Leng, Y., Bolton, M.D. 2012. Construction of hairpin RNA expressing vectors for RNA-mediated gene silencing in fungi. In: Bolton, M.D., Thomma, B.P.H.J. editors. Plant Fungal Pathogens: Methods and Protocols. New York, NY: Humana Press. p. 623-633.
Bolton, M.D., Birla, K., Rivera-Varas, V., Rudolph, K.D., Secor, G.A. 2012. Characterization of CbCyp51 from field isolates of Cercospora beticola. Phytopathology. 102(3):298-305.
Vargas, M., Loyola, C., Zapata, N., Rivera, V., Secor, G., Bolton, M.D., France, A. 2012. First report of root rot of Chicory caused by Phytophthora cryptogea in Chile. Plant Disease. 96(4):591.
Dejonge, R., vanEsse, P., Maruthachalam, K., Bolton, M.D., Santhanam, P., Saber, M.K., Zhang, Z., Usami, T., Lievens, B., Subbarao, K.V., Thomma, B.P. 2012. Tomato immune receptor Ve1 recognizes effector of multiple fungal pathogens uncovered by genome and RNA sequencing. Proceedings of the National Academy of Sciences. 109(13):5110-5115.
Nelson, B.D., Bolton, M.D., Lopez-Nicora, H.D., Niblack, T.L., Del Rio Mendoza, L. 2012. First confirmed report of sugar beet cyst nematode, Heterodera schachtii, in North Dakota. Plant Disease. 96(5):772.