Location: Temperate Tree Fruit and Vegetable Research
2020 Annual Report
Objectives
Develop or identify new breeding lines, germplasm and named cultivars with superior quality, disease and pest resistance, and nutritional value. This will involve collaborative and independent work by our three-person team using our respective expertise in potato breeding, molecular physiology and plant pathology. The three objectives below undertake complimentary approaches to germplasm improvement. Objective 1 involves largely breeding for targeted traits. Objective 2 seeks to determine basic mechanisms that govern trait expression. Objective 3 will develop new or improved methods to evaluate breeding lines and germplasm. We will work closely with the TriState Breeding Program, as we have for over 20 years.
Objective 1: Evaluate, identify, breed, and release potato germplasm with improved traits of interest, especially improved disease and pest resistance, and increased amounts of phytonutrients.
Subobjective 1A. Develop breeding lines, cultivars or identify germplasm with enhanced amounts of phytonutrients and visual appeal.
Subobjective 1B. Develop breeding lines, cultivars or identify germplasm with superior disease resistance with a focus on soil-borne diseases.
Objective 2: Characterize genetic, environmental, molecular, physiological, and biochemical factors that control accumulation of potato phytonutrients and mechanisms that lead to plant disease resistance, and use this knowledge to produce new superior potato cultivars.
Subobjective 2A: Determine mechanisms that mediate tuber phytonutrient expression.
Subobjective 2B: Increase information and develop methods with potential to be used for control of Potato Cyst Nematode (PCN) and for improved disease resistance.
Objective 3: Develop improved pathogen diagnostic techniques and phenotyping approaches that can be used for potato germplasm evaluation, development of host-resistance, and identification of emerging potato diseases.
Subobjective 3A. Identify and characterize emerging and evolving pathogens and pests in the Pacific Northwest.
Subobjective 3B: Characterize Tobacco rattle virus (TRV)-potato interactions to develop better detection methods and determine the relationship between viral titer, cultivar, symptoms and resistance.
Approach
1A. Germplasm will be intercrossed and progeny evaluated in the field. Replicated plots will be grown in successive years across multiple locations. Lines will be analyzed for carotenoids, anthocyanins, antioxidants, total protein, potassium and iron. Molecular markers will be used to characterize high carotenoid lines. Liquid chromatography mass spectrometry (LC-MS) will be used to quantitate phytonutrients. If germplasm does not provide the desired traits, we will import additional germplasm.
1B. Resistance to nematodes, viruses and fungi will be developed using resistant lines to make crosses and evaluating progeny in field trials. Selected clones will be evaluated under high disease pressure and molecular markers used for Meloidogyne chitwoodi breeding. If progeny have lower selection rates than expected the size of the initial population will be increased.
2A. Expression of structural genes and transcription factors in potatoes or organs that have low or high amounts of phenolics, are cold-treated, or wounded will be analyzed using reverse transcription quantitative polymerase chain reaction (RT-qPCR) and LCMS. We will use ribonucleic acid sequencing (RNA-seq) to generate transcriptomic data. Effect of environment on glycoalkaloids will be assessed by growing 13 genotypes in six locations and methanolic extracts from freeze-dried tubers analyzed by LCMS. If key genes are identified, resources will be redirected to apply this knowledge through precision breeding efforts.
2B. Potato cyst nematode (PCN) trap crop seed will be produced by sowing true seed directly into the soil at ¼ inch depth. Hatching factor purification will be tested on diverse High-performance liquid chromatography (HPLC) columns and fractions tested for activity. If a hatching factor is identified and quantitated by LCMS, increased resources will be directed.
3A. Samples from symptomatic plants will be collected. Grafting experiments will evaluate transmissibility. Established molecular tools will be used to detect any pathogens present. If targeting known pathogens does not identify a biological agent, primers that target unknown pathogens will be used. Psyllid involvement in beet-leafhopper transmitted virescence agent (BLTVA) will be tested using field and cage experiments. Development of improved diagnostic tools for BLTVA and Candidatus Liberibacter solanacearum (Lso) will be assessed using a single-tube nested PCR technique, RT-qPCR or Kompetitive allele-specific PCR. If unable to identify any known pathogen in a sample, next generation sequencing platforms will be used.
3B. Tobacco Rattle Virus (TRV) sampling methods will be evaluated for efficacy. Lines will be evaluated for resistance in field trials. PCR will be used to compare viral titer with symptom severity. Varieties will be exposed to TRV and differences in resistance/insensitivity and susceptibility compared. Daughter tuber symptoms and viral titer will be compared to mother tuber symptoms, viral titer, plant emergence, and daughter tuber yield. If TRV infection becomes sporadic, we will focus on the genotypes that were subject to sufficient disease pressure.
Progress Report
In support of Sub-objectives 1A, 1B, and 2A scientists in Prosser, Washington, conducted a potato crossing block in the spring of 2020 generating thousands of recombinant seeds to be evaluated in future field trials and acquired both genotype and disease symptom data that is being used to develop a molecular marker to select for Tobacco rattle virus (TRV) resistance in progeny from Castle Russet. Progress was made by introgressing extreme resistance to viral diseases (Potato Virus Y (PVY), TRV, Potato mop top virus (PMTV)) and nematode infection (Columbia root knot nematode (CRKN), Potato cyst nematodes) into susceptible genetic backgrounds. Progeny material containing the Rysto allele (PVY immunity) can be selected using molecular genotyping techniques and those that inherit resistance will be sent for field selection. Hybridizations performed with material confirmed to be CRKN resistant will set up a progeny test panel to identify resistant clone and elite parent combinations that exhibit superior general and specific combining ability for yield and processing traits like specific gravity. Breeding lines were analyzed for nutritional value, focusing on compounds called phenylpropanoids that have numerous health-promoting effects, including anti-inflammation and anti-cancer.
For Sub-objectives 1A, 1B, and 2A, scientists in Prosser researched potato glycoalkaloids. Glycoalkaloids influence potato disease resistance and nutritional value. Crosses were made between high and low glycoalkaloid lines to develop a population that can be used to identify genes that control glycoalkaloid amounts. Transgenic potatoes with increased and decreased amounts of sterols were made to assess the effect on tuber greening and glycoalkaloid amounts. Expression of genes involved in glycoalkaloid metabolism were measured. Light treatments were administered to whole tubers to assess the effect on greening, glycoalkaloids, and gene expression in response to light. Studies were conducted to assess how flavonol biosynthesis is regulated in potatoes. Flavonols have numerous health-promoting benefits and are dietarily desirable but are present in low amounts in tubers. We identified transcription factors that regulate flavonols and showed a regulatory loop between sucrose and transcription factors. We demonstrated that transcription factors that regulate potato phenylpropanoid metabolism bind the promoters of genes that degrade sucrose.
Baby potatoes are a growing market and among the best options for the potato industry to target emerging consumer preferences, such as nutrition, taste, novelty, ease of preparation, and visual appeal. One limitation to the development of this market is reduced yields, which are far less than when potatoes are grown to maturity. In support of Sub-objective 1A and 2A scientists in Prosser, Washington, have developed a promising breeding line that continued to produce exceptionally high numbers of tubers in another year of field trials, consistent with its performance in previous years. Because of its unusual, highly-desirable characteristics, we used this line to make numerous additional crosses in an attempt to fast-track development of a high-tuber set potato and transfer the high-set trait into additional germplasm and accelerate the breeding process that normally takes over 12 years.
In support of Sub-objective 1B, and 3A scientists in Prosser, Washington, completed pathogen inoculation study designed to identify the mechanism of corky ringspot resistance present in Castle Russet. Findings indicate that Castle Russet is immune to TRV infection but can still act as a host to the stubby root nematodes that are vectors. A mock trial project to identify physical and biological factors associated with soil health status as it relates to corky ringspot and CRKN was completed. In support of Sub-objective 2B scientists in Prosser, Washington, evaluated the agronomic performance of Litchi Tomato, a potential trap crop for the potato cyst nematode. Additionally, these plants were evaluated for potato viroids in collaboration with APHIS.
In support of Sub-objectives 1A and 1B scientists in Prosser, Washington, developed proximal and remote sensing technology to assess tuber characteristics, plant health, and environmental variables associated with disease symptoms. In collaboration with scientists in Aberdeen, Idaho, a semi-automated data acquisition workflow to measure tuber size, shape, and colorimetric features of tuber skin and flesh was developed. Values extracted from digital images were found to be highly correlated with manual measurements of tuber length and width acquired using digital calipers. Aerial orthomosaic images of experimental plots at four different time points throughout the growing season (seedling emergence, row closure, flowering, and harvest) were acquired. A trait extraction pipeline was constructed, and results showed differences in both plant size and reflectance characteristics between tobacco rattle virus infested and unaffected fields. The ability of hyperspectral sensors to discriminate between plant genotypes, medium, and TRV infection status was evaluated.
For Sub-objectives 3A and 3B scientists in Prosser, Washington, and North Dakota State University examined if TRV-insensitivities are due to environmental, nematode, or TRV population differences between North Dakota and Washington States. Two potato cultivars showing possible insensitivity to TRV tuber symptom development and one cultivar that is sensitive to the virus-induced symptom development were inoculated with nematodes harboring TRV that originated from North Dakota or from Washington State. At harvest, nematode populations were quantified from the soil, and potato foliage, roots, and tubers were tested for TRV. In all treatments, the number of nematodes was reduced with the North Dakota population compared to the Washington population. Despite this, for the sensitive cultivar, the percentage of TRV infected tissue was higher with the North Dakota nematode population than the Washington population. These results indicate that biological differences may exist between the two nematode populations, which could result in different transmission efficiencies or rates of TRV infection. Disease severity index scores collected from tubers in 2019 and 2020 suggest that the TRV immunity locus encoded by Castle Russet is inherited as a dominant, single locus trait and preliminary linkage mapping results indicate that this trait is controlled by a major quantitative trait locus (QTL) on the distal end of chromosome IX. In 2020, we planted this population in our corky ringspot evaluation field and assessed yield, specific gravity, tuber number, size, shape, color and disease symptoms. Progress was made on the optimization of a reverse transcription quantitative-polymerase chain reaction method for assessing viral titer of TRV in tubers.
In support of Sub-objective 3A, scientists in Prosser, Washington, examined the role potato psyllid may have in the acquisition and transmission of the beet leafhopper transmitted virescence agent (BLTVA) phytoplasma in potato. Three greenhouse experiments were completed that forced potato psyllids to feed on BLTVA-infected potato plants. Potato psyllids were subsequently recovered from the plants and tested for the presence of the pathogen by molecular diagnostics. In these controlled experiments, psyllids acquired BLTVA at very low levels despite being caged on infected plants. In these controlled experiments, infection levels were lower than those that have been identified in natural field-caught potato psyllids. Collaboration with scientists at Wapato, Washington, is underway to utilize a fluorescent labeling technique to visualize the presence of the pathogen in insects under the microscope. We hope to compare pathogen presence and localization in the known BLTVA vector, the beet leafhopper, with the potato psyllid. Differences in pathogen location within each insect will validate the results of our greenhouse studies.
In support of Sub-objective 3A, scientists in Prosser, Washington, identified the insect vector of the novel ‘Candidatus Liberibacter solanacearum’ (Lso) haplotype F bacterium that causes internal defects in tubers. In collaboration with Oregon State University scientists, psyllids were collected from the Klamath Basin of Oregon from sticky cards placed in or near commercial potato fields. Scientists in Prosser, Washington, then tested over 1200 and 1500 psyllid specimens from these traps in 2018 and 2019, respectively. Psyllids were generally identified as belonging to the Aphalara genus through visual identification technique and molecular analysis. A subset of psyllids tested positive for the Lso pathogen in both 2018 and 2019, and work to identify the specific haplotype of the bacterium is nearing completion. To help growers in the Klamath Basin potato growing region prevent the spread of disease caused by Lso haplotype F, it is critical that the insect vector is identified to determine the best pest management strategies to implement moving forward.
Additionally in support of Sub-objective 3A, scientists in Prosser, Washington, performed diagnostic analysis to determine the presence of two important soilborne viruses in potato seed lots planted by commercial growers in the Columbia Basin of Washington and Oregon were performed. Results identified a low, but persistent level of the economically important Potato mop top virus in seed lots originating from states across the United States and in Canada. Infected seed lots consist of different russet and specialty potato cultivars. This work has demonstrated that seed can be a source of the virus entering commercial fields, but work is still needed to determine if the virus-free vector can pick up the virus from infected seed.
Accomplishments
1. Discovery of a mechanistic link between potato greening and glycoalkaloid production. Greening of potatoes can cause various issues related to health, grower profitability, increased waste and trade issues. Tuber greening is caused by an increase in chlorophyll, but the issue comes from a concomitant increase in unrelated toxic compounds called glycoalkaloids. Using a molecular approach, ARS scientists at Prosser, Washington, and Washington State University identified a mechanism that links greening and glycoalkaloid biosynthesis for the first time. Manipulating the general isoprenoid pathway altered chlorophyll and glycoalkaloid biosynthesis, and also influenced the amount of health-promoting carotenoids and increased biomass. These findings create new approaches to develop potatoes resistant to greening and increase crop productivity.
2. Corky ringspot resistance encoded within Castle Russet is due to immunity to Tobacco rattle virus. Corky ringspot is a widespread, soil-borne potato tuber disease caused by Tobacco rattle virus infection and vectored by stubby root nematodes that feed on plant roots. Corky ringspot is currently managed through the application of soil fumigants to reduce the abundance of the nematode vector, but once affected by corky ringspot, field remediation is extremely difficult. By combining pathogen inoculation assays in a controlled environment and molecular diagnostics, ARS scientists in Prosser, Washington, determined that the mechanism of genetic resistance to corky ringspot observed in Castle Russet is due to immunity to Tobacco rattle virus and not resistance to the stubby root nematode vector. Evidence from this study demonstrates that viral presence in diseased soils can be reduced or eliminated within four months if the only food source accessible to the nematode vector is Castle Russet or alfalfa. This discovery suggests that cultivation of Castle Russet in rotation with other Tobacco rattle virus resistant crops like alfalfa may help growers maintain profitability while remediating corky ringspot affected fields by reducing viral titer.
3. Seasonal population dynamics of the economically damaging/devastating potato psyllid evaluated, documented, or identified in the Columbia Basin potato growing region of Washington and Oregon. Potato psyllid is a potential economically devastating pest of potato and other solanaceous crops in the United States that growers must control each year through integrated pest management strategies that are often very costly. Predicting potato psyllid population levels prior to their arrival in potato crops could be very beneficial to growers seeking to prevent excessive use of costly insecticides for both environmental and economic reasons. Researchers in Wapato and Prosser, Washington, in collaboration with Washington State University scientists sought to determine how seasonal winter and summer weather patterns might affect psyllid population dynamics and the presence of an economically devastating pathogen that the psyllid transmits to potato. In addition to length of cold temperatures during winter, data suggest that degree-day accumulation over time and total degree-day accumulation affect psyllid population size each year. Psyllid infection rate with the pathogen ranged from 0-0.17% in the four-year study, indicating that pathogen levels are low in this region. This research indicates that winter conditions, degree-day accumulation, and time of first catch are indicators growers could use to adjust their pest management strategies on a yearly basis to likely be more cost- and time-effective.
4. Computer vision data acquisition and measurement workflow established to quantify size, shape, and colorimetric characteristics of potato tubers. Tuber size, shape, skin, and flesh quality are a few of many factors assessed by the food processing industry when they consider accepting a new potato variety. Reproducible and accurate measurement of these characteristics on thousands of tubers generated in each breeding and evaluation cycle is needed to apply statistical genetics models and marker assisted selection on these traits within a potato breeding program. Scientists in Prosser, Washington, and Aberdeen, Idaho, developed a semi-automated data acquisition workflow to measure these features using a microcomputer, consumer grade digital single-lens reflex camera (DSLR), and flatbed scanner. Tuber size, shape, and the colorimetric features of tuber skin and flesh were extracted from digital images and found to be highly correlational to manual measurements of tuber length and width acquired using digital calipers. The capability to inexpensively and reproducibly quantify many tuber traits simultaneously on the volume of samples encountered in breeding populations (thousands per year) provides increased value to our stakeholders. These tools and the datasets collected using these techniques can be easily stored, shared with collaborators, and reused for different purposes such as the development of machine learning models to classify and quantify tuber defects.
Review Publications
Jansky, S.H., Navarre, D.A., Bamberg, J.B. 2019. Introduction to the special issue on the nutritional value of potato. American Journal of Potato Research. 96:95-97. https://doi.org/10.1007/s12230-018-09708-1.
Si, M., Lange, I., Lange, B.M., Navarre, D.A. 2020. Effect of HMGR and FPS overexpression and silencing on potato sterol synthesis and growth. Journal of Experimental Botany. 71(14):4109-4124.
Bibi, S., Navarre, D.A., Sun, X., Du, M., Rasco, B., Zhu, M. 2019. Beneficial effect of potato consumption on gut microbiota and intestinal epithelial health. American Journal of Potato Research. 96:170-176. https://doi.org/10.1007/s12230-018-09706-3.
Ellsworth, P.Z., Feldman, M.J., Baxter, I.R., Cousins, A.B. 2020. A genetic link between leaf carbon isotope composition and whole-plant water use efficiency in the C4 grass setaria. Plant Journal. 102(6):1234-1248. https://doi.org/10.1111/tpj.14696.
Swisher Grimm, K.D., Mustafa, T., Cooper, W.R., Munyaneza, J.E. 2020. Growth and yield performance of Solanum tuberosum grown from seed potatoes infected with ‘Candidatus Liberibacter solanacearum’ haplotypes A and B. Plant Disease. 104:688-693. https://doi.org/10.1094/PDIS-05-19-1125-RE.
