Research Project: Genetic Improvement of Blackberry, Red and Black Raspberry, Blueberry, and Strawberry

Location: Horticultural Crops Production and Genetic Improvement Research Unit

2020 Annual Report

Objectives
Objective 1: Develop new improved small fruit cultivars for the commercial fruit industry, including, thornless, machine harvestable blackberry cultivars with excellent flavor; high yielding, virus tolerant, high quality strawberry cultivars with outstanding processing characteristics; virus tolerant, high yielding, black raspberry selections with flavor and color characteristics similar to the current standard cultivars; and high quality primocane fruiting red raspberry cultivars with broad adaptation. Objective 2: Evaluate, and incorporate new sources of genetic variability from Rubus, Vaccinium, Fragaria and other minor crops. Specifically, selections of Rubus occidentalis populations from across the country will be evaluated for fruit quality, plant growth, and aphid resistance, and strawberry populations for the presence of repeat flowering characteristics. • Subobjective 2A: Incorporate new/novel germplasm into breeding material to enhance adaptation. • Subobjective 2B: Determine whether trait-associated markers can be developed and if developed, can they be used to identify germplasm that expresses a phenotype of interest. Objective 3: Characterize viruses that infect berry crops, and develop management strategies to minimize the impact of virus diseases on these crops. • Subobjective 3A: Sequence novel viruses of small fruit crops and develop assays for their rapid detection, emphasis will be on raspberry leaf curl disease and two novel virus-like diseases of grapevine. • Subobjective 3B: Determine if endogenous Rubus yellow net virus (RYNV) in raspberry genomes pose a risk of long distance virus spread. • Subobjective 3C: Eradicate Blueberry fruit drop associated virus from blueberry fields in the U.S. Objective 4: Develop virus-tested planting stocks of berry cultivars for fruit production using thermal therapy and other virus elimination techniques, and test RNAi therapy as a new method for virus elimination in berry crops.

Approach
For each crop, a modified recurrent mass selection system will be used. Individuals that are selected in a given generation will be intercrossed to produce the next generation. For Rubus spp. and strawberry 4000-6000 seedlings, and 2500-4000 blueberry seedlings from 30-100 crosses will be evaluated annually. Approximately 0.5 to 1.0% of the seedlings are selected based on subjectively evaluated fruit quality, plant performance, ripening period, and yield. Frozen samples will be evaluated for processing characteristics. Selections identified as being superior will be propagated for commercial trial and release. For nursery production, the ARS virology program in Corvallis, OR will help produce virus-tested propagation material. To broaden the germplasm base, superior individuals or representatives of superior populations of small fruit species will be crossed among themselves or with advanced selections or cultivars. Selections from these crosses will be used in our breeding program and distributed to other breeders. Emphasis will be on aphid resistance, disease resistance, fruit quality, thornlessness and tolerance to abiotic stresses associated with growing these crops in regions with more diverse climates. This program collaborates with molecular geneticists in efforts to bridge the gap between genomics and applied plant breeding in berry crops. Our program helps to determine the mapping populations to develop or genotypes to include and how the various phenotypic traits (phenological, reproductive, and vegetative) will be evaluated. The phenotyping for each project is being coordinated across multiple locations with different climatic conditions. For raspberry leaf curl disease, suspect raspberry samples will be collected and their virome analyzed using Next Generation Sequencing. This approach will also be used to examine two novel diseases of grapevines in Oregon. Diagnostic assays will be developed and used for epidemiology studies, certification and quarantine purposes. Virus vectors will be identified and strategies for vector control developed as a means to manage virus diseases. Total genomic sequencing of five Rubus cultivars will be used to determine if the inserted RYNV sequences represent full-length or partial RYNV and if all insertions are at the same site. If the RYNV insertions are at the same site in each of these cultivars it would indicate that the insertion may have happened once and been passed on in breeding programs. Several approaches will be tested for applying gene silencing as a tool to eliminate viruses from growing points of plants (meristematic dome and several leaf primordia): 1. Virus specific dsRNA will be produced and provided to plants in tissue culture as an additive to the media; 2. dsRNA will be attached to positively charged clay nanoparticles and sprayed on plants in tissue culture or growth chambers; and 3. RBDV infected plants will be grafted onto transgenic plants (already developed red raspberry) that are producing RNAi silencing of RBDV. Meristems will be collected at various times after treatments, plants regenerated and tested for RBDV.

Progress Report
There are currently no SYs on this project. The lead scientist/berry breeder passed away in December, 2019 and the other berry breeder on this project retired in December, 2019. However, activities still continue in both of these research programs until these critical vacancies are filled. Towards Objective 1, the “Celestial Blackberry Series” was released from the USDA/Oregon State University cooperative breeding program. They are all thornless, semi-erect and high quality blackberry releases. ’Galaxy’ blackberry has large fruit. ‘Twilight’ is suited for the fresh market and ripens in the early mid-season. ‘Eclipse’ has firm, dark fruit of uniform shape well suited for the fresh market. Patent data continues to be collected from advanced blueberry and raspberry selections that the former lead scientist indicated were worthy of patenting. For Objective 2, in the berry breeding program (blueberry, caneberry, and strawberry), seedlings from crosses made in 2019 are being evaluated. Advanced selections are being maintained and harvested to determine growth characteristics and fruit quality. For Objective 3, viral high-throughput sequence data has been archived to support future research in novel virus discovery, small RNAs and to continue characterizing raspberry leaf curl disease. ARS researchers in Corvallis, Oregon, continue work on improving virus elimination efficiency for developing ‘clean plants’ as part of the National Clean Plant Network. The test viruses for this project are Raspberry bushy dwarf virus and Strawberry necrotic shock virus, which are the most difficult viruses to eliminate using standard thermal therapy treatments. Research on the use of small interfering RNAs (RNAi) to reduce the quantity of virus in a given volume (virus titer) in plants is being tested as a foliar application and as an additive to tissue culture media. In the initial studies with RNAi, the virus titer was reduced by as much as 90%. In parallel experiments, RNAi is being combined with thermal therapy or chemotherapy, and chemotherapy is being combined with thermal therapy. Meristems have been taken from the RNAi treated plants and are being grown out for further analysis.

Accomplishments

Review Publications
Vance, A., Jones, P., Finn, C.E., Strik, B. 2019. Fruit development in blackberry types and cultivars – Impact of days and temperature from bloom to stages of ripening. Journal of American Pomological Society. 73(4):227-239.
Finn, C.E., Strik, B.C., Yorgey, B.M., Peterson, M.E., Jones, P.A., Buller, G., Serce, S., Lee, J., Bassil, N.V., Martin, R.R. 2020. ‘Eclipse’ thornless semi-erect blackberry. HortScience. 55(5):749-754. https://doi.org/10.21273/HORTSCI14891-20.
Finn, C.E., Strik, B., Yorgey, B.M., Peterson, M.E., Jones, P.A., Lee, J., Bassil, N.V., Martin, R.R. 2020. 'Twilight' thornless semi-erect blackberry. HortScience. 55(7):1148-1152. https://doi.org/10.21273/HORTSCI14992-20.
Bradish, C., Bushakra, J., Robbins, L., Karaaoac, E., Sabrina, T., Willard, J.L., Perkins-Veazie, P., Lee, J., Scheerens, J., Weber, C., Dossett, M., Bassil, N.V., Finn, C.E., Fernandez, G. 2020. Standardized phenotyping in black raspberry. Journal of American Pomological Society. 74(1):2-17.
Finn, C.E., Strik, B., Yorgey, B.M., Peterson, M.E., Jones, P.A., Buller, G., Lee, J., Bassil, N.V., Martin, R.R. 2020. ‘Galaxy’ thornless semierect blackberry. HortScience. 55(6):967-971. https://doi.org/10.21273/HORTSCI14985-20.
Sooriyapathirana, S., Ranaweera, L., Perera, H., Weebadde, C., Finn, C.E., Bassil, N.V., Hancock, J.F. 2019. Using SNP/INDEL diversity patterns to identify a core group of genotypes from FVC11, a superior hybrid family of Fragaria virginiana Miller and F. chiloensis (L.) Miller. Genetic Resources and Crop Evolution. 66:1691–1698. https://doi.org/10.1007/s10722-019-00819-0.
Tzanetakis, I.E., Martin, R.R. 2019. Improving plant propagation methods for fruit disease control. In: Xu, X., Fountain, M., editors. Integrated Management of Diseases and Insect Pests of Tree Fruit. Cambridge, UK: Burleigh Dodds Science Publishing. p. 275-288. http://dx.doi.org/10.19103/AS.2019.0046.13.
Diaz-Lara, A., Martin, R.R., Rwahnih, M.A., Vargas, O.L., Rebollar-Alviter, A. 2019. First evidence of viruses infecting berries in Mexico. Journal of Plant Pathology. 102:183–189. https://doi.org/10.1007/s42161-019-00381-9.
Zurn, J.D., Ivors, K.L., Cole, G.S., Knapp, S.J., Hummer, K.E., Hancock, J.F., Finn, C.E., Bassil, N.V. 2020. Assessing cultivated strawberries and the Fragaria Supercore for resistance to soilborne pathogens. Journal of American Pomological Society. 74(1):18-23.
Worthington, M.I., Aryal, R., Bassil, N.V., Mead, D., Fernandez, G.E., Clark, J.R., Fernandez-Fernandez, F., Finn, C.E., Hummer, K.E., Ashrafi, H. 2020. Development of new genomic resources and tools for molecular breeding in blackberry. Acta Horticulturae. 1277:39-46. https://doi.org/10.17660/ActaHortic.2020.1277.6.
Willman, M., Bushakra, J., Bassil, N.V., Finn, C.E., Dossett, M., Fernandez, G., Weber, C., Scheerens, J., Dunlap, L., Fresnedo-Ramirez, J. 2020. Genetic analysis of drupelet count in black raspberry (Rubus occidentalis). Acta Horticulturae. 1277:65-72. https://doi.org/10.17660/ActaHortic.2020.1277.9.
Zurn, J.D., Meiers, R.C., Ward, J., Finn, C.E., Dossett, M., Bassil, N.V. 2020. Identifying variation in red raspberry MLO genes thought to provide resistance to powdery mildew. Acta Horticulturae. 1277:25-32. https://doi.org/10.17660/ActaHortic.2020.1277.4.
Bushakra, J., Alice, L., Carter, K., Dossett, M., Lee, J.C., Liston, A., Meiers, R., Mulch, C., Nyberg, A.M., Peterson, M.E., Clark, M.C., Vining, K., Worthington, M., Yin, M., Sutherland, B., Zurn, J.D., Clark, J., Finn, C.E., Bassil, N.V., Hummer, K.E. 2020. Status of Rubus germplasm at the US National Clonal Germplasm Repository in Corvallis, Oregon. Acta Horticulturae. 1277:121-128. https://doi.org/10.17660/ActaHortic.2020.1277.17.