Research Project: Genetic Improvement of Lettuce, Spinach, Celery, Melon, and Related Species

Location: Crop Improvement and Protection Research

2023 Annual Report

Objectives
Objective 1: Develop genetic resources and germplasm resistance to key pests and pathogens and abiotic stresses of lettuce, spinach, celery, and melon. Sub-objective 1.A: Breed for resistances to fungal, bacterial, and viral diseases and disorders in lettuce. Sub-objective 1.B: Develop durable resistance to downy mildew in spinach. Sub-objective 1.C: Improve celery for resistance to Fusarium oxysporum f.sp. apii (Foa) races 2 and 4. Sub-objective 1.D: Breed melon for resistance against powdery mildew, virus, and whitefly. Objective 2: Develop new procedures and technologies to evaluate and enhance postharvest quality and shelf life of fresh cut leafy greens, especially lettuce. Objective 3: Improve content and bioavailability of phytonutrients or fiber of leafy vegetables for improved impact on human health nutrition and composition of the gut microbiome. Sub-objective 3.A: Characterize polyphenol and fiber content among lettuce cultivars. Sub-objective 3.B: Breed red spinach for improved nutrient content and disease resistance. Objective 4: Characterize the composition and effects of environment on the microbiota found on leafy vegetables.

Approach
Objective 1: Sub-objective 1A: Map major QTLs for resistance to downy mildew using a genome-wide association mapping (GWAS) approach and develop breeding lines with the improved resistance to lettuce drop and downy mildew. Characterize resistance to Fusarium, Verticillium, and Pythium wilts to develop improved lettuce germplasm. Cross corky root-resistant variety ‘Glacier’ and wild species L. serriola to sources of other disease resistance and select for combined resistances to corky root, leafminer, downy mildew, lettuce mosaic virus, tipburn, and horticultural and nutritional traits in different types of lettuce. Characterize resistance to impatiens necrotic spot virus to develop improved lettuce germplasm. Develop controlled environment and molecular assays to characterize tipburn resistance in lettuce. Sub-objective 1B: Breed open-pollinated (OP) spinach with resistance to all prevalent downy mildew races through crosses in isolators, recurrent selection, and replicated field trials. Sub-objective 1C: Develop celery germplasm with resistance to Foa races 2 and 4 through disease assays, selection, and self-pollination. Sub-objective 1D: Breed western U.S. shipper type melon resistant to powdery mildew races 1, 2, 3.5, 5, and S. Identify and characterize resistance in melon to whitefly-transmitted cucurbit chlorotic yellows virus (CCYV). Characterize antixenosis to sweetpotato B biotype whitefly in melon. Objective 2: Fine-map the locus for slow decay of fresh-cut lettuce, map QTLs for limited oxidative browning, and develop lettuce breeding lines with high postharvest quality. Objective 3: Sub-objective 3A: Characterize polyphenol and fiber content among lettuce cultivars to understand genotype x environment interactions and to identify genetically stable high polyphenol cultivars in replicated greenhouse assays. Sub-objective 3B: Improve the betacyanin content, antioxidant capacity, and downy mildew resistance of different types of spinach through crossing, recurrent selection, and field trials. Objective 4: Determine the relationships in the phyllosphere community among leafy vegetables, nearby weeds, and the soil microbiota by isolating genomic materials from the phyllosphere and rhizosphere for sequencing.

Progress Report
This report documents progress for project 2038-21530-003-000D, Genetic Improvement of Lettuce, Spinach, Celery, Melon, and Related Species, which started April 2023 and continues research from project 2038-21530-002-000D, Genetics and Breeding of Lettuce, Spinach, Melon, and Related Species to Improve Production and Consumer-related Traits. In support of Objective 1, Sub-objective 1.A, Goals 1.A.1 and 1.A.2, seeds of 500 lettuce accessions were planted in a field to produce seeds for all subsequent experiments. In support of Sub-objective 1.A, Goal 1.A.3, an ARS researcher at Salinas, California, conducted a greenhouse test in May to evaluate commercial lettuce varieties and internationally recognized Fusarium differential varieties to characterize reports of “new races” in commercial fields. Germplasm was inoculated with multiple Fusarium isolates representing race 1 and the new variant. Discussions with ARS and university plant pathologists is leading to coordinated efforts for biological and molecular characterization of these and other isolates and a clear direction for resistance breeding. Researchers at Salinas, California, planted a field trial in June to evaluate USDA breeding lines and check varieties under Verticillium race 1. Selections for disease resistance and horticultural traits will be conducted in September and promising lines will be moved forward for germplasm development and release. In support of Sub-objective 1.A, Goal 1.A.4, we are analyzing data from our field trials of advanced generations for resistances to corky root, leafminers, downy mildew, tipburn, and horticultural traits in different types of lettuce. In support of Sub-objective 1.A, Goal 1.A.5, researchers at Salinas, California, conducted a greenhouse test in May to evaluate a mapping population of 162 recombinant inbred lines (RILs), parents, and check varieties for resistance to impatiens necrotic spot virus (INSV). Results will identify regions of the genome linked to resistance alleles of interest for breeding and germplasm development and release. Researchers at Salinas, California, will plant fields trials in June and August to evaluate commercial varieties, USDA breeding lines, and check varieties for resistance to INSV. We will harvest in early fiscal year (FY) 2024. In support of Sub-objective 1.A, Goal 1.A.6, research continued to develop a controlled environment to characterize tipburn resistance. Two trials in a growth room at low growing temperatures and high water content has produced tipburn in some susceptible cultivars. More replication is necessary to confirm the method is reproducible. In support of Sub-objectives 1.B and 3.B, we are harvesting spinach seeds from isolators for the recurrent selection of downy mildew resistance and betacyanin content. Harvested seeds are being cleaned for planting in next year’s selection nursery. In support of Sub-objective 1.C, research continued on celery resistance to Fusarium oxysporum f.sp. apii (Foa) races 2 and 4. Crosses were made of Foa race 2-resistant germplasm (cv. Challenger) and Foa race 4-resistant germplasm (cv. French Dinant and University of California, Davis germplasm lines) to develop germplasm with stacked resistance to Foa race 2 and 4. Crosses were also made between Foa race 4-susceptible (cv. Challenger) and Foa race 4-resistant germplasm (USDA PI181714 or A0134) to develop a mapping population to identify genomic regions associated with resistance. In support of Sub-objective 1.D, four F2:F3 populations were generated for identifying Quantitative Trait Loci (QTL) for resistance to powdery mildew (PI 313970 x Top Mark) and cucurbit yellow stunting disorder virus (CYSDV; PI 313970 x Top Mark), and sweetpotato whitefly (PI 313970 x Top Mark, PI 122847 x Top Mark, TGR 1551 x Top Mark, and PI 414723 x Top Mark). Two QTL for CYSDV resistance were identified, one of them was expressed in two seasons and located in linkage group 5, and two potential markers for marker assisted selection were identified. Identification of QTL for resistances powdery mildew races 1 and 2 are underway. Inheritance of resistance and QTL analysis for whitefly resistance is underway. A triple septet of powdery mildew differentials was established, increased and deposited in USDA NPGS (U.S. National Plant Germplasm System) for distribution. A melon core collection/set for genetic analyses and screening of economically important traits was established. The set consists of 384 diverse melons (from North America, Europe, Africa, Central and East Asia and India) and represents approximately 99% of the genetic diversity in approximately 2083 melon accessions mostly contained in the USDA, NPGS melon collection. The set is in the process of being made more uniform via a process of single seed descent for submission to USDA NPGS. In support of Objective 2, seeds of recombinant inbred lines (RIL) with genetic differences in the qSL4 locus were planted in a greenhouse. The resulting plants will be used for crossing to develop a population of genotypes for fine mapping qSL4, the gene controlling lettuce postharvest quality. In support of Sub-objective 3.A, research continued on polyphenol and fiber content among lettuce cultivars. Between 48-52 cultivars of lettuce were grown in a growth room for 6-8 weeks in two trials, and samples will be processed to assay total polyphenol content. A subset will also be processed for total fiber content. In support of Objective 4, substantial planning has been completed to characterize the composition and effects of environment on the microbiota found on leafy vegetables. Sites have been identified for collections, and collections will take place later in the summer.

Accomplishments

Review Publications
Bhattarai, G., Shi, A., Mou, B., Correll, J. 2023. Skim resequencing finely maps the downy mildew resistance loci RPF2 and RPF3 in spinach cultivars Whale and Lazio. Horticulture Research. 10(6). Article uhad076. https://doi.org/10.1093/hr/uhad076.
Simko, I., Hasegawa, D.K., Peng, H., Zhao, R.B. 2023. Genetic and physiological determinants of lettuce partial resistance to Impatiens necrotic spot virus. Frontiers in Plant Science. 14. Article 1163683. https://doi.org/10.3389/fpls.2023.1163683.
Peng, H., Simko, I. 2023. Extending lettuce shelf life through integrated technologies. Current Opinion in Biotechnology. 81. Article 102951. https://doi.org/10.1016/j.copbio.2023.102951.