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.
This is the final report for project 8080-21000-022-00D which terminated in March 2018. Below is a summary of the progress over the life of the project. Breeding efficiency can be enhanced by the use of DNA tests for important traits. Fire blight is an economically important disease of apple and breeding for resistance to this disease is a priority in several apple breeding programs.‘Splendour’ apple is both fire blight resistant and of superior fruit quality making it an excellent parent in breeding for resistance. Based upon genetic analysis it was determined that the fire blight resistance of ‘Splendour’ was due to the interaction of three genetic loci on chromosomes 4, 5 and 11. DNA markers were developed for these loci and are being validated on known fire blight resistant and susceptible ‘Splendour’ progeny from multiple crosses. Once validated, these DNA tests will provide a valuable tool for plant breeders to efficiently breed fire blight resistant apple cultivars. Durable disease resistance which lasts for many years is particularly important in apple due to the long production period of apple orchards. A common approach to enhance the durability of a disease resistant cultivar is to pyramid many different resistance genes for a single disease within the cultivar. To facilitate pyramiding resistance for fire blight disease the National Plant Germplasm System’s collection of apple cultivars and wild species accessions are being screened for novel fire blight resistance genes in collaboration with Cornell University. In order to identify novel fire blight resistance variants, 304 apple accessions known to be highly resistant to fire blight are being screened for resistance with multiple strains of the fire blight pathogen and being screened with DNA tests for 5 previously identified fire blight resistance genes. Information on apple accessions with novel fire blight resistance genes will be made available to the wider apple research community and industry through GRIN-Globaldatabase to facilitate durable fire blight resistance through resistance gene pyramiding. Fire blight is a devastating bacterial disease of apple trees that results in severe economic losses. It is known that a specific protein produced by the fire blight bacteria, called AvrRpt2EA, is important for pathogen recognition in the fire blight resistant crabapple ‘Robusta 5’. However, little is known about the role of this protein in susceptible apple varieties. In collaborative studies with the Julius Kuhn Institute of Germany the AvrRpt2EA protein was expressed in fire blight susceptible ‘Pinova’ apple plants using a heat-inducible gene expression system in order to study the function of this protein. After the AvrRpt2EA protein was expressed in susceptible apple plants they developed symptoms similar to natural fire blight infections with browning leaves and death of vegetative shoots. Expression of the protein also resulted in increased levels of the plant hormone salicylic acid (SA) and the systemic acquired disease resistance pathway it controls. This work demonstrates that AvrRpt2EA protein acts as virulence factor and plays an important role in the antagonism between SA- and JA-signaling to facilitate the further colonization of the infected plant by the fire blight pathogen. Host plant resistance is one of the most effective and sustainable options for managing fire blight, a devastating disease of apple and pear. Since genetic diversity is often lost during crop domestication, accessions of Malus sieversii, the wild progenitor of the domesticated apple, represent a valuable resource for disease resistance. Nearly 200 accessions of Malus sieversii from a USDA-ARS collection were selected as potential sources of disease resistance for apple scion breeding. Based upon controlled challenges with the fire blight pathogen, 12 wild Malus sieversii accessions were identified as resistant to fire blight with resistance comparable to highly resistant Malus × robusta 5 and resistant 'Delicious'. Three of these M. sieversii accessions PI657116, GMAL3688.c and GMAL4002.k were crossed in FY2017 with ‘Pinova’ (PinataTM) to begin incorporating this resistance into apple breeding programs. Developing frost protection methods for flowering fruit trees during spring frost events is critical. Transgenic MDCBFx plants with appropriate controls were planted as indicated in the milestone goals for FY2018. An inter-graft experiment with CBF-overexpressing scions is nearing completion to examine whether light perception is important for graft-transmission of altered dormancy. A root chilling experiment was conducted to assess whether maintaining root temperatures = 40 °F would delay bud break. Per the milestone goals for FY2018, apple trees over expressing the two apple versions of the EBB (Early Bud Break) gene have been created along with trees over expressing the poplar EBB1 gene; trees are growing in the greenhouse and are being evaluated.
Desnoues, E., Norelli, J.L., Aldwindkle, H.S., Wisniewski, M.E., Evans, K.M., Malnoy, M., Khan, A.M. 2018. Identification of novel strain-specific and environment-dependent minor QTLs linked to fire blight resistance in apples. Plant Molecular Biology Reporter. 36:247-256. https://doi.org/10.1007/s11105-018-1076-0.
Dong, C., Ma, Y., Wisniewski, M.E., Cheng, Z. 2017. Meta-analysis of the effect of overexpression of CBF/DREB family genes on drought stress response. Environmental and Experimental Botany. 142:1-14.
Dong, C., Ma, Y., Zheng, D., Wisniewski, M.E., Cheng, Z. 2018. Meta-analysis of the effect of overexpression of dehydration-responsive element binding family genes on temperature stress tolerance and related responses. Frontiers in Plant Science. https://doi.org/10.3389/fpls.2018.00713.
Harshman, J., Evans, K., Allen, H., Potts, R.J., Flamenco, J.A., Asldwinckle, H.S., Wisniewski, M.E., Norelli, J.L. 2017. Fire blight resistance in wild accessions of Malus sieversii. Plant Disease. 101(10):1738-1745.
Ballester, A., Norelli, J.L., Burchard, E.A., Abdelfattah, A., Levin, E., Gonzalez-Candelas, L., Droby, S., Wisniewski, M.E. 2017. Transcriptomic response of resistant (P161983-Malus sieversii) and susceptible ("Royal Gala") genotypes of apple to blue mold (penicillium expansum) infection. Frontiers in Plant Science. 8(1981):1-16.
Hamdoun, S., Gao, M., Gill, M., Kwon, A., Norelli, J.L., Lu, H. 2017. Signalling requirements for Erwinia amylovora-induced disease resistance, callose deposition and cell growth in the non-host Arabidopsis thaliana. Molecular Plant Pathology. 19(5):1090-1103. https://doi.org/10.1111/mpp.12588.
Levin, E., Ballester, A., Raphael, G., Feigenberg, O., Liu, Y., Norelli, J.L., Gonzalez-Candelas, L., Ma, J., Dardick, C.D., Wisniewski, M.E., Droby, S. 2017. Identification and characterization of LysM effectors in Penicillium expansum. PLoS One. 12(10):1-25.