Location: Fruit and Tree Nut Research
2024 Annual Report
Objectives
1. Improve pecan nut productivity by analyzing key horticultural issues that disrupt annual consistency, yield, and quality, and developing new or improved mitigation strategies. [NP305, C1, PS1B] (Previously reviewed and approved)
1.A. Determine if canopy exposure to nano-particles, in particular zinc and nickel nano-particles can improve health and longevity of pecan tree canopies.
1.B. Characterize horticultural traits of native pecan germplasm and identify genes of interest as a resource for development of new and improved cultivars.
1.C. Characterization of improved pecan rootstocks for uniformity of yield and enhanced productivity.
2. Reduce impacts of the most important pecan diseases on production, quality and uniformity of nutmeats. [NP305, C1, PS1B] (Previously reviewed and approved)
2.A. Characterize and identify novel ways to improve management of pecan scab in tree canopies based on inoculum sources, fungicide spray coverage, disease distribution and methods for disease management.
2.B. Determine dynamics of population genetic diversity of Venturia effusa in pecan orchards.
3. Develop new and improved pecan processing technologies, such as pasteurization and cracking/shelling, for improved storage, food safety, nutrition, and marketability. (NP305, C1 PS1B; NP 306, C1, PS1C) (New, for review)
3.A. Determine factors that influence the growth and/or survival of foodborne pathogens on the surface of whole and cracked pecans.
3.B. Evaluate the efficacy of novel technologies as mitigation strategies to reduce foodborne pathogens on pecans.
3.C. Identify, characterize, and model pecan and machine factors to improve halves yields in pecan cracking and shelling.
Approach
This research aims to provide pecan farmers with improved, sustainable tree and disease management practices that stabilize yield in pecan (Carya illinoinensis) orchards and maximize postharvest nutmeat quantity and quality. The management tools and strategies will enable farmers to mitigate alternate bearing (AB) and yield loss caused by disease. AB is considered the most important biological problem facing pecan production: it is economically harmful, resulting in excessive year-to-year fluctuation in nut yield and kernel quality. Many biotic and abiotic factors can induce or increase the amplitude of AB. How factors associated with canopy health, particularly nutrition, rootstock, and disease affect AB represent some of the knowledge gaps that limit development of suitable tools for stabilizing nut production and reducing yield losses. Losses postharvest include physically damaged kernels due to postharvest processing, loss to biological contaminants and poor shelf life resulting in loss of quality and salability. The research addresses 1. Whether use of nano-fertilizers can provide a basis for more efficient nutrient management, stabilize and increase production of pecan, while ensuring better environmental security. 2. Phenotyping horticultural traits of native and improved pecan germplasm in conjunction with genome wide analyses. 3. Using genomics to identify markers for horticultural traits for use in the breeding program. 4. Characterizing the role of rootstocks in tree growth and productivity, to provide a basis for more uniform, consistent and thus sustainable production of pecan nutmeats. 5. Determining how inoculum sources of pecan scab (caused by Venturia effusa) contribute to the epidemic, and using this information to develop new disease management tools, thereby reducing the impact of scab in susceptible pecan cultivars. 6. Understanding the population genetics of the scab pathogen to underpin deployment of more durable host resistance in the future. 7. Using genomics to identify effector genes involved in scab pathogen virulence. 8. Developing novel nut shelling and associated processing techniques to minimize physical nutmeat damage, contamination, and loss of quality postharvest. and 9. Improving understanding of pre-harvest and postharvest environmental factors that impact pecan color, texture, oil quality, and phytochemical content to identify conditions that maximize duration of quality. To better understand food safety risks associated with pecans, the project will address factors that influence the survival and persistence of foodborne pathogens and identify effective mitigation strategies while maintaining nut quality. In order to maximize marketability, the project will explore means to cost-effectively increase intact pecan meat extraction by modifying parametrized process variables on existing processing equipment. A series of field and laboratory studies over the project duration will address the key knowledge gaps and provide greater knowledge and understanding to develop horticultural and food safety tools and strategies required for improved management and thus long-term sustainability of pecan.
Progress Report
ARS researchers at Byron, Georgia collected data of agronomically and horticulturally important traits from a pecan collection. Characterized bud break and flowering time in all trees in the provenance collection housed at ARS, Byron, Georgia. Additional tree phenotypic traits including trunk diameter, tree height, leaf area, nut load and tree architecture were collected and are being analyzed. Nut quality traits are being assessed. Collected the phenotypic data for fruit developmental period, date of fertilization was recorded from all trees. Date of shuck-split will be recorded when fruit matures. Period between fertilization and shuck-split will provide information on fruit developmental period. Collected leaf material from the collection and extracted DNA for sequencing to conduct genome wide association studies for SNP identification. The traits are important for future breeding programs and will contribute to improved, sustainable production and will benefit the health value of the pecan nut.
A fourth year’s collection of tree phenotypic variables was made in the ‘Lakota x 87MX3-2.11’ mapping population. Variables measured included growth rate (tree height, trunk diameter), leaf area, and bud break. All transplants in the mapping population were assessed for leaf scab (recording the percentage area scabbed on the most severely diseased leaflet and providing an overall disease score for the plant). Data will be analyzed for QTL mapping. The trees are still young and will provide more comprehensive data in coming years. The trial is repeated in Texas. The two different climates in Texas and Georgia will allow environment x genotype studies, providing a basis for understanding inheritance of different phenotypic traits in pecan.
Agronomic and horticultural traits were assessed in pecan trees grafted on hickory rootstocks. Variables included tree height, trunk diameter, nut load, yield, and nut quality characteristics. The cultivars Desirable and Stuart were grafted on 12 species of hickory at the Southeastern Fruit and Tree Nut Research Station, Byron, Georgia in 1986 to establish graft compatibility. The aim now is to determine whether different species of hickory rootstock influences tree yield and nut quality of the two common pecan cultivars. The study will provide information on whether non-pecan hickories may be beneficial rootstocks to improve pecan production.
Nuts were harvested over the developmental period of nut growth including water, shell hardening, and kernel fill stages from cultivars Elliott, Pawnee, and Sumner. Allergen analysis of proteins, including RNA activity is currently in progress with the collaborator in Albany, California. This time course study will allow for a better understanding of pecan nut attributes such as lipid content, lipid oxidation, and allergens during pecan nut growth and development.
A new trial was set up at Shiloh Farms in Ray City, GA looking at the effect of the plant growth regulator ReTain (Valent USA, LLC) on nut retention. During the first season, nut load was assessed early, mid, and late season post application. Trees were hedged in 2022 and assessments occurred on hedged vs non-hedged sides of trees. At harvest, yield (in shell and total nutmeat) was obtained. Data collected included nut quality parameters and a second year data collection is underway. This trial will allow us to assess whether the product will lower the incidence of nut drop on cv. Desirable while following regular hedging management strategies.
A new pecan block of cultivars Avalon and Lakota was successfully established. Trees that did not survive the first year’s planting were replaced and are being established. The block comprises 328 trees planted on a 10 x 10 m spacing and is equipped with below-surface drip irrigation. This new planting will allow for studies on nutrient and irrigation management, future hedge-pruning, and other research on two new scab-tolerant cultivars that were otherwise not planted.
Scab severity was assessed for a fourth year in a mapping population designed to better understand the inheritance of scab resistance. The cross is the two genotypes ‘Lakota’ x ‘87MX3-2.11’. Assessments were performed for each genotype in the experiment. As part of a multi-year, multi-location study, the information will aid understanding of scab resistance for breeding purposes.
An experiment to assess effects of tractor travel speed and spray application volume was conducted. Building on prior experiments application volumes of 114, 470 and 940 L/ha and speeds 2.4 and 3.2 km/h were compared. Samples of leaves and fruit were taken at three different heights in the canopies of the trees, and speed and volume effect on disease control explored. If fungicides can be applied at faster travel speeds and lower volumes without reducing efficacy, growers can save substantial time and other operational cost.
Laboratory experiments to ascertain effects of different phosphite fungicide formulations on growth and spore production of V. effusa were conducted. There is little detailed information on the effect of alternative phosphites (with Ni, Cu, Mn or Zn) on the growth of the fungus, but they may be more efficacious than the standard K formulation. Improved efficacy will result in better control of scab disease.
A second years’ experiment to study scab lesions on shoots was conducted. Shoot lesions are considered to be a primary source of disease early in the season. Determinate, fruiting shoots and indeterminate shoots were collected, and the numbers of lesions counted at the end of the season. Spore production was also quantified. Better knowledge of shoot lesions and their role in the disease cycle should provide a basis for improved control.
Functional elements and genome architectures were characterized in the annotated reference genomes of three scab pathogens (Venturia effusa, V. nashicola, and V. inaequalis) to investigate alternate splicing events and intron structure of scab pre-mRNA’s. Sequence attributes of Venturia pre-mRNA’s are conserved across multiple scab species including the 5’ untranslated region, Kozak sequence, splice site donor/acceptor, polypyrimidine and uridine tracts, as well as the branchpoint nucleotide. RNA-sequencing data, harvested from the NCBI sequence read archives, is currently being processed for novel transcript isoform discovery in three scab pathogen species. Predictions of the branchpoint motif using a position weight matrix (in-progress) will deconvolute alternate splicing events arising from intron retention and exon skipping modules. Intron classes are being designated by covariate features including rank, length, A/T/C/G frequencies, splice donor motif, splice acceptor motif, branchpoint motif, and branchpoint distance from 3’ intron terminus. Together, these data will elucidate alternatively spliced isoforms of Venturia transcriptomes, and they demarcate cis-regulatory sequences involved in the regulation of scab gene expression.
Work started on validating the plant hormone ethylene as playing a major role in susceptibility to pecan scab. RNA was extracted from half-sib cultivars Elliott and Desirable exogenously treated with ethylene. Quantitative PCR was run to check the expression pattern of six different genes predicted to be responsive to ethylene. Leaves of various cultivars of pecan were also checked for ethylene generation via gas chromatography at University of Georgia in Athens. Half-sibs of Elliott and Desirable were also treated with ethylene and inoculated with Venturia effusa conidia to check for increased susceptibility to the fungal pathogen.
Plasmids (pRN119 & pRN120) expressing developmental regulator genes were obtained, transformed into Agrobacterium tumefaciens, and used for “in soil” transformation techniques. Theses plasmids have been shown to increase transformation efficiency in plants. Several phloem specific promoters have been identified and sub-cloning work started for future translating ribosomal affinity purification sequencing (TRAP-seq) experiments of pecan. Due to Venturia effusa’s affinity to vascular tissue, transcripts specific to this tissue could reveal important susceptibility factors.
Mislabeled plants in a previously unused genome wide association dataset have been identified and removed for more accurate results of downstream applications. A pipeline for population structure and metagenomic analysis of this dataset, and future datasets, was developed. Using population structure information generated from this dataset a tree from the Provenance collection with unique genetics compared to historical cultivars was identified that also has high resistance to pecan scab. Genetic crosses were started between this individual and susceptible cv Desirable trees for ease assessing genetic contributions to scab resistance/susceptibility. Markers for identifying successful crosses have been identified and await development. Microbiome analysis was also done using this dataset and shows unique stratification of micro-organisms associating with pecan compared to previous microbiome studies. In conjunction with past scab severity ratings, this information could be used to develop biopesticides.
A pilot study was started emasculating cv Cape Fear branches to check for either improved or detrimental effects to nut development/yield. The provenance collection was also scouted for individuals producing few catkins. Future work using breeding, biotechnology techniques, or amendment application to produce female only trees could reduce alternate bearing and/or increase yield.
Putative genetically transformed Venturia effusa (the organisms causing pecan scab) are awaiting materials/resources required for characterization of their genetics.
The Pecan Processing Aspects of the project have been separated and documented in the report for project 6042-41000-001-000-D.
Accomplishments
1. Visual assessments of pecan canopy density due to late season leaf fall provided accurate and reliable estimates for comparing pecan genotypes.. Canopy density during leaf fall is important as the seasonal canopy dynamics provides insights to environmental cues and breeding potential of germplasm. ARS researchers at Byron, Georgia explored the ability of visual raters to estimate canopy foliage density in trees in the provenance collection as an indicator of leaf shed during autumn. The estimates were compared to measurements using image analysis. The results are a basis for understanding genetic control of leaf fall when combined with genotype data as part of a genome-wide association study.
2. Analysis of transcriptomes from pecan trees with wild ancestry reveal potential mechanisms of resistance/susceptibility to pecan scab.. Pecan scab disease is a key factor that limits pecan productivity in most of the eastern US growing region. ARS researchers at Byron, Georgia collected RNA from pecan trees previously shown to be either susceptible or resistant to pecan scab. Infected leaf material in addition to healthy material was collected from susceptible trees. Next-gen sequencing revealed transcriptome profiles of these different materials. Pathway enrichment analysis of this information revealed potential candidates of resistance and/or susceptibility. These genes and their associated pathways are being analyzed in further detail to guide breeding efforts for enhanced resistance to pecan scab.
Review Publications
Pisani, C., Bock, C.H., Randall, J. 2023. Visual rating and the use of image analysis for assessing canopy foliage density in a pecan provenance collection during leaf fall. Journal of Forestry Research. 34:1843-1834. https://doi.org/10.1007/s11676-023-01635-0.
Toledo, P., Phillips, K., Schmidt, J., Bock, C.H., Wong, C.R., Hudson, W., Shapiro Ilan, D.I., Wells, L., Acebes-Doria, A.L. 2023. Canopy hedge pruning in pecan production differentially affects groups of arthropod pests and associated natural enemies. Crop Protection. 176. Article 106521. https://doi.org/10.1016/j.cropro.2023.106521.
Brungardt, J.J., Alarcon, Y., Shiller, J., Monteros, M., Young, C., Randall, J.J., Bock, C.H. 2024. Transcriptome profile of pecan scab resistant and susceptible trees from a pecan provenance collection. BMC Genomics. 25:180. https://doi.org/10.1186/s12864-024-10010-0.
Chiang, K., Chang, Y., Liu, H., Lee, J., Jarroudi, M., Bock, C.H. 2024. Survival analysis as a basis to test hypotheses when using quantitative ordinal scale disease severity data. Phytopathology. 114(2). https://doi.org/10.1094/PHYTO-02-23-0055-R.
Johnson, K.A., Brannen, P.M., Chen, C., Bock, C.H. 2024. Visual assessment of phony peach disease: evaluating rater accuracy and reliability. Plant Disease. 108(4):930-940. https://doi.org/10.1094/PDIS-11-22-2669-RE.
Johnson, K.A., Chen, C., Bock, C.H., Brannen, P.M. 2023. No evidence for vertical transmission of Xylella fastidiosa subsp. multiplex in peach seed. Plant Health Progress. 24(4):516-518. https://doi.org/10.1094/PHP-11-22-0121-BR.
Pisani, C., Cottrell, T.E. 2024. Impact of Pecan Leafroll Mite (Acari: Eriophyidae) on Pecan Foliage. Journal of Entomological Science. 59(1):86-89. https://doi.org/10.18474/0749-8004-59.1.86.