Plant Pathologist

Professional Biographical Information:

Ph.D. in Plant Pathology (1986), Cornell University

M.S. in Plant Pathology (1978), University of California, Berkeley

B.S. in Agricultural Science (1973), Cornell University

2001-present; Research Plant Pathologist; USDA-ARS, Appalachian Fruit Research Station, Kearneysville, WV.

1978-2001; Senior Research Associate (1993-2001), Research Associate (1987-1993), Research Support Specialist (1978-1986); Cornell University, New York State Agricultural Experiment Station, Geneva, NY.

Description of Research Projects:

My long-term research goals are to characterize the underlying basis of disease resistance in fruit trees and to develop novel control strategies for specific tree-fruit diseases.  Fire blight of apple, caused by the bacterial pathogen Erwinia amylovora, is the focus of the research. The aim of the research is to enhance economic opportunities for agricultural producers (ARS Strategic Goal 1) by fostering the development of new fire blight control technologies, thereby reducing the threat of this devastating disease to the U.S. fruit industry and decreasing the industry’s cost of production; and to enhance protection and safety of the nation’s agriculture and food supply (ARS Strategic Goal 3) by mitigating concerns regarding the commercial use of genetically enhanced fruit crops.

Current research topics include:

Genomic response of apple to fire blight infection:

Goal: Use functional genomics to characterize the response of apple to fire blight disease and, thereby, identify new opportunities for improving fire blight resistance.

Approach: Global approaches are being used to characterize changes in apple gene expression in response to fire blight challenge. A fire blight-susceptible and –resistant apple cultivar are challenged with the fire blight pathogen, E. amylovora, and the transcriptome (all expressed genes) of the two cultivars is characterized by cDNA-AFLP and cDNA hybridization subtraction. Subtractive cDNA hybridization is also being used to investigate the basis for susceptibility by challenging the susceptible cultivar with an incompatible pathogen, E. amylovora and defined mutants of the pathogen altered in their ability to secrete ‘effector’ proteins (that are believed to elicit disease development) into host cells. The genes are identified by genomic profiling and there function in fire blight susceptibility or resistance will be studied by reducing or knocking-out expression of the gene, and then observing its effect on phenotype and gene expression. To accomplish this we are developing a high-throughput system for generating RNA interference (RNAi) mutants in apple using ESTs associated with fire blight. The goal of a high-throughput approach is to generate a large number of mutants from an unselected population of candidate genes to allow large-scale reverse genetic analysis of genes associated with a specific trait.

Collaborators:
Dr. Herb Aldwinckle, Professor of Plant Pathology, Cornell University, Geneva, NY
Dr. Carole Bassett, Molecular Biologist, USDA-ARS, Kearneysville, WV
Dr. Robert Farrell, Assistant Professor of Biology, Penn. State Univ., York, PA
Dr. Timothy McNellis, Assistant Professor of Plant Pathology, Penn. State Univ., University Park, PA
Dr. Michael Wisniewski, Research Plant Physiologist, USDA-ARS, Kearneysville, WV

Development of new technologies to manage fire blight:

Goal: Improve management of fire blight with new control technologies that enhance host resistance.

Approach: Although increasing host resistance has been recognized as an important component of fire blight management, its application has been limited by a lack of resistant cultivars suited to commercial needs and by a lack of management practices that could effectively increase resistance. Recent advances have made it feasible to change this paradigm in the 21^st century. First, apple rootstock breeding programs have developed dwarfing rootstocks that are resistant to fire blight and are currently becoming available for commercial use (Norelli et al. 2003b). Second, the feasibility of genetically engineering commercial apple cultivars for increased fire blight resistance has been demonstrated and transgenic apple plants are now undergoing field trials (Norelli et al. 2003a). Third, chemical treatments that enhance host resistance have been demonstrated to be useful in the control of fire blight (Norelli and Miller 2004, Norelli et al. 2003a). New technologies to enhance host resistance in fruit crops will continue to be developed as our understanding of host resistance mechanisms is advanced through research.

Trait modification through genetically enhanced rootstocks:

Goals: Mitigate safety concerns regarding the use of transgenic apple varieties by developing technology to use genetically enhanced rootstocks to deliver trait modification to conventional (i.e. not genetically engineered) apple scions.

Background: Genetically enhanced, or engineered, plants have the potential to improve crop performance, improve food quality and increase farm profits. However, there are many hurdles to be overcome before genetically enhanced apples can become a reality in the orchard. Among the most important hurdles are consumer concerns regarding the safety of genetically engineered plants and the impact this may have on marketing. There are also significant environmental and commercial concerns regarding the unintended spread of transgenic pollen to wild and homeowner apple plants, to nearby organic blocks or to blocks targeted for export that could result in serious economic losses.
The application of rootstock mediate trait modification could overcome many of the hurdles facing genetically enhanced (sometimes referred to as transgenic) fruits. These include eliminating the risk of transgenic pollen spread since pollen would not be produced by the genetically engineered rootstock, improved consumer acceptance since fruit will not be transgenic, and greater ease of commercialization since a single rootstock could be used to enhance the value of many different commercial fruiting varieties.

Approach: Genetic enhancement a specific trait in a rootstock will not automatically result in trait modification of the scion; specific biological mechanisms will need to be employed in the rootstock to make the trait modification graft-transmissible. We are investigating the use of gene silencing and the expression of transgenes in the vascular tissue of the rootstock as two possible ways to accomplish graft-transmissible trait modification.
Gene silencing in transgenic plants has been demonstrated to be an effective method to develop plants with novel traits. Examples of how this technology is being used in fruit trees include the development of virus and fire blight resistance, fruit with delayed softening, and trees with altered architecture. Because gene-silencing does not involve the introduction of “foreign” genes but rather “switches off” existing genes, it is considered a “soft” transgenic technology. In tobacco, gene silencing has been shown to be graft-transmissible from transgenic rootstock to non-transformed scions. Although the targeted trait in the scion is altered, the transgene is not present in the scion and it is not genetically modified. Graft-transmissible gene silencing has not been adequately investigated in transgenic fruit trees to predict how it will function in apple. However, it has a high likelihood of being a useful mechanism to facilitate rootstock mediated trait modification in apple scions.
Although gene silencing promises to be a useful mechanism for trait modification, it can not be used for all types of trait modification. Some trait modifications will require the activity of altered or new proteins in the scion cultivar. In these cases, the rootstock will need to be engineered to deliver the new protein to the conventional scion by means of plant’s vascular system. This could be accomplished by expressing the protein in the vascular tissue of the rootstock thus allowing the protein to move from the rootstock to the scion. Although this approach will be technically more difficult, basic research in tobacco and other model systems indicate that it should be possible in apple by targeting expression of the transgene to specific cells in the rootstock.

Collaborators:
Dr. Carole Bassett, Molecular Biologist, USDA-ARS, Kearneysville, WV
Dr. LaiLiang Cheng, Assist. Professor of Horticulture, Cornell Univ., Ithaca, NY
Dr. Gennaro Fazio, Plant Geneticist, USDA-ARS, Geneva, NY
Dr. Michael Wisniewski, Plant Physiologist, USDA-ARS, Kearneysville, WV

Infection of apple shoots by Erwinia amylovora (fire blight):

Goal: Identify the sources of fire blight bacteria that initiate the shoot blight phase of the disease.

Background: Although significant progress has been made in the last 20 years in our ability to predict and control the blossom blight phase of fire blight, the shoot blight phase of the disease remains poorly understood and difficult to control. One of the major gaps in our understanding of shoot blight is the origin or source of the bacteria that cause shoot infection. Epiphytic bacteria, that is bacteria multiplying on the plant surface without causing disease, are known to be important in the blossom blight phase of the disease and predicting the build up of these bacteria in blossoms is a critical component to predicting blossom blight infection periods. However, it is not known if E. amylovora can actively multiply on apple shoots.

Approach: The initial goal of this study was to determine if E. amylovora could multiply and survive on apple leaves as an epiphyte. We were unable to demonstrate the epiphytic multiplication of E. amylovora on the leaf surface based upon inoculation under controlled environmental conditions, orchard monitoring and microscopic observation. However, we were able to detect foliar populations of E. amylovora under orchard conditions that occurred following rapid temperature changes during summer storms.
Recent results suggest that wetting events during high temperatures can lead to the colonization of hydathodes and glandular trichomes and the establishment of E. amylovora within young leaves. Under controlled conditions, E. amylovora did not multiply epiphytically on leaves at constant 24^o C. However, when plants were inoculated with cold bacteria (4^o C) and incubated at high temperature (35^o C), E. amylovora quickly became established within young leaves but rapidly declined on the surface of older leaves. Microscopic observation of an E. amylovora strain labeled with green fluorescent protein (gfp) indicated that under these conditions (cold inoculum, high post-inoculation incubation temperature), bacteria would colonize hydathodes and glandular trichomes of young leaves.

Collaborators:
Dr. Maria Brandl, Microbiologist, USDA-ARS, Albany, CA

The role of carbohydrates metabolism in fire blight resistance:

Goals: Determine if the accumulation of sorbitol in apple tissue inhibits the growth of E. amylovora, and thereby increases the plants resistance to fire blight.

Background: Based upon observed sorbitol concentration in apple tissue, osmotic potential and shoot susceptibility to infection by E. amylovora, Suleman and Steiner hypothesized that increases in sorbitol concentration result in increasingly negative solute potentials that have negative effects on the growth of E. amylovora, thus rendering these tissues more resistant (Phytopathology 84:1244-1250. 1994).

Approach: A genetic approach is being used to test the Suleman/Steiner hypothesis by down-regulating the genes encoding the rate limiting enzymes for the synthesis of sorbitol and sucrose in apple, NADP-sorbitol-6-phosphate dehydrogenase (S6PDH) and sucrose phosphate synthase. Apples and other Rosaceous plants accumulate photoassimalates as sucrose, sorbitol and starch. Down regulating S6PDH in apple results in lower sorbitol concentration but has little effect on osmotic potential due to higher sucrose content in these plants. Down regulation of these enzymes will be accomplished by transforming apple with RNAi constructs.
In addition, “biosensors” are being developed to quantify both E. amylovora perception of osmotic stress and its utilization of sorbitol in apple tissue during the infection process. To accomplish this, an E. amylovora DNA promoter that responds to osmotic stress is being fused to a reporter gene that can be quantified in apple tissue. Similarly, sorbitol utilization will be quantified using the srlA promoter that has been reported to be induced in E. amylovora by sorbitol under both in vitro and in planta conditions.

Collaborators:
Dr. LaiLiang Cheng, Assist. Professor of Horticulture, Cornell Univ., Ithaca, NY