1: Improve environmental stress and disease resistance in tree fruit crops. 1.A. Identify and characterize sources of fire blight resistance for use in apple scion breeding programs. 1.B. Characterize expression patterns and sequence differences of select apple drought-responsive genes in Malus sieversii lines exhibiting high and low water use efficiency. 1.C. Utilize transcriptomic and high-throughput genetic screening approaches to identify CBF-regulated and other stress-regulated genes, and characterize their functional role in stress tolerance and dormancy using transgenics and field evaluation. 2: Develop an accelerated breeding system for new tree fruit crops utilizing transgenic early-flowering lines.
Abiotic and biotic stresses play a major role in determining the economic viability of fruit crop production and postharvest quality. A single fire blight epidemic can destroy an entire young orchard and unfavorable environmental conditions, such as freezing temperatures, as well as heat and drought stress can result in significant reductions in yield, quality, and tree longevity. The overall objective of this project is to utilize genomic and molecular approaches to identify genes that convey resistance to abiotic and biotic stress in fruit crops, identify genetic markers for disease and stress resistance that can be utilized by apple breeders in marker-assisted-breeding programs, and to develop a breeding system that will facilitate the incorporation of specific traits, especially from novel genetic resources, such as Malus sieversii, into advanced selections of breeding lines. Quantitative trail loci (QTLs) and molecular markers for fire blight resistance will be developed for resistance derived from Malus sieversii and ‘Splendour’ apple. Targeted genome sequencing of the promoters of select dehydration and water use efficient responsive genes will be applied to lines of xeric-adapted Malus sieversii. Promoter analysis will identify cis-elements known to affect gene expression. Methylation differences between lines during simulated drought will be evaluated to reveal potential targets for gene regulation. The contribution of the CBF (C-repeat binding factor) family members to cold hardiness, dormancy, and growth will be evaluated. An accelerated breeding system for apple will be developed utilizing transgenic early-flowering lines to facilitate rapid integration of important genetic traits from novel apple genotypes into advanced breeding lines. The proposed research will result in the identification of genes, molecular markers, and a breeding system that can be used to efficiently develop apple germplasm with increased resistance to biotic and abiotic stress.
Host plant resistance is one of the most effective and sustainable options for managing fire blight, a devastating disease of apple and pear. 'Splendour' apple, which has excellent flavor and is resistant to fire blight, was found to transmit its resistance to 25% of its progeny and will, therefore, be a useful donor of resistance for apple breeding programs. In order to facilitate the identification of specific genetic markers for resistance, the fire blight resistance of 600 'Splendour' progeny was determined in controlled evaluation trials. Climate change has resulted in early budbreak in many species of fruit crops making them very susceptible to subsequent freezing events. Therefore, there is a need to develop cultivars or management practices that allow trees to adapt to these changes in climate in order to maintain yields. A model system has been developed utilizing transgenic lines of apple that overexpress CBF genes in order to understand how CBF regulates growth, dormancy, and cold hardiness in fruit trees. This work has been published. Understanding how ice is initiated and how it propagates in plants can lead to the identification of new approaches for frost protection. Protocols have been published that demonstrate the utility of using high-resolution infrared thermography to study ice nucleation and propagation in plants. The use of this technology by USDA-ARS has revolutionized how frost tolerance is studied and evaluated in plants. Identifying genes expressed in cold hardy, dormant trees vs. those that are less hardy and lose dormancy early will help to better understand how these processes are regulated in fruit trees. Transcriptomic studies of gene expression in wild-type and transgenic apples overexpressing several different CBF transcription factor genes have been completed and data is being analyzed. Developing genetic markers for postharvest disease resistance can facilitate breeding programs that utilize marker-assisted selection. Crosses have been made between early flowering lines and genotypes of a Malus sieversii x 'Royal Gala' cross in order to determine if the fruit produced in the offspring are resistant and if specific markers are present. This effort is being made to validate the markers that have been associated with postharvest resistance to blue mold. Breeding new apple varieties takes 25-30 years due to its long juvenile phase. Rapid cycling breeding using "early flowering" transgenic apple has the potential to greatly accelerate apple breeding. Crosses of early flowering 'Pinova' T1190 with fire blight and apple scab resistant 'Enterprise' and 'Goldrush' resulted in the production of 250 early flowering seedling that are being selected for disease resistance.
1. A model system to study the impact of stress-tolerance genes on the physiology of apple (Malus X domestica). Regulating dormancy and frost tolerance in apple is essential as warmer winters and spring followed by episodes of freezing weather are causing greater and greater economic losses to fruit growers. ARS researchers in Kearneysville, West Virginia, constructed a transgenic apple in which the overexpression of a transcription factor results in early and prolonged dormancy, and increased freezing tolerance. This model system is now being used to better understand how processes such as dormancy, growth, and freezing tolerance are interconnected and regulated together in a synchronous manner. This knowledge is required to develop effective breeding, management, and transgenic approaches to increasing spring frost tolerance without impacting growth and yield.
2. Fire blight resistance identified in the wild large-fruited progenitor of domesticated apple (Malus sieversii). Fire blight of apple and pear is estimated to cost American fruit growers over $1M per year in lost production and control. Although genetic resistance is considered the most efficient and reliable means of controlling this devastating disease, there is a lack of resistance in domesticated apple. ARS researchers in Kearneysville, West Virginia, identified 12 accessions of wild Malus sieversii with excellent fire blight resistance that were originally collected by ARS in Kazakhstan. Unlike other wild apple species, M. sieversii is the only wild species with large edible fruit, making it especially well suited for breeding new apple cultivar with resistance to fire blight.
Bassett, C.L., Artlip, T.S., Wisniewski, M.E. 2015. Water deficit treatment and measurement in apple trees. Bio-protocol. 5(3):1-7.
Wisniewski, M.E., Norelli, J.L., Artlip, T.S. 2015. Overexpression of a peach CBF gene in apple: a model for understanding the integration of growth, dormancy, and cold hardiness in woody plants. Frontiers in Plant Science. DOI: 10.3389/fpls.2015.00085.
Buron-Moles, G., Wisniewski, M.E., Vinas, I., Teixido, N., Usall, J., Droby, S., Torres, R. 2015. Characterizing the proteome and oxi-proteome of apple in response to a compatible (P. expansum) and a non-host (P. digitatum) pathogen. Journal of Proteomics. 114:136-151.
Wisniewski, M.E., Neuner, G., Gusta, L.V. 2015. The use of high-resolution infrared thermography (HRIT) for the study of ice nucleation and ice propagation in plants. Journal of Visualized Experiments. DOI: 10.3791/52703.
Ballester, A., Marcet-Houben, M., Levin, E., Sela, N., Selma-Lazaro, C., Carmona, L., Wisniewski, M.E., Droby, S., Gonzalez-Candelas, L., Gabaldon, T. 2015. Genome, transcriptome, and functional analyses of Penicillium expansum provide new insights into secondary metabolism and pathogenicity. Molecular Plant-Microbe Interactions. 28(3):232-248.