INTEGRATED GENOMICS-ENABLED BREEDING FOR ACCELERATING SOIL-BORNE DISEASE RESISTANCE IN STRAWBERRY (F. X ANANASSA)

Fresh strawberries are a vital component of the U.S. fruit and vegetable industry, with an annual farm gate value of approximately $2.5 billion. Economic losses due to root and crown rots caused by soilborne pathogens are estimated at $150 million annually in the U.S. In the Florida strawberry industry alone, yield losses due to soilborne diseases are estimated tocost growers $15 million in revenue each year, despite annual pre-plant soil fumigation. Phytophthora crown rot (PhCR), caused by Phytophthora cactorum, and charcoal rot (CR), caused by Macrophomina phaseolina, are currently the most damaging and widespread soilborne diseases of strawberry in the U.S. and the world, causing up to 40% production losses. Crop mortality can surpass 80% in CR-infected fields of susceptible cultivars. Limitations on fumigants such as methyl bromide have led to deterioration in the control of P. cactorum and M. phaseolina in recent years. Registration of new fumigants has been difficult, and alternatives to methyl bromide have had lower and inconsistent efficacy for a cultivar of pathogens. Commercial cultivars vary widely in their genetic resistance to these soilborne pathogens. The cultivars most resistant to PhCR provide sufficient levels of disease control, but currently, the cultivars most resistant to CR provide only moderate disease reduction. Thus, the development of resistant cultivars is becoming a top priority for strawberry breeding programs, and a critical strategy for combatting these diseases in an era of alternative fumigants. Breeding efforts and genomics research at the University of Florida have identified large-effect QTL for resistance to both PhCR and CR under field conditions. For PhCR, the resistance locus is located on chromosome 7B and is referred to as FaRPc2. Within the FaRPc2 locus, two significant haplotypes, FaRPc2-H2 and FaRPc2-H3, confer resistance against P. cactorum. Each resistant haplotype explained about 40% of the phenotypic variance in the tested populations. The resistant haplotype, FaRPc2-H3, is more prevalent in commercial strawberry cultivars in the United States. The two haplotypes originated from different sources, suggesting the presence of two distinct functional alleles associated with the FaRPc2-mediated resistance. Cultivars with FaRPc2-H2 display strong resistance to PhCR; however, genomic tools to investigate FaRPc2-H2 are missing, which is a bottleneck for deploying genomics-assisted breeding strategies. For CR resistance, our group recently discovered three large-effect resistance QTL on chromosomes 2B (FaRMp1), 4A (FaRMp2), and 4C (FaRMp3). Each locus explained 20% to 40% of the phenotypic variance in the tested populations. While FaRMp1 and FaRMp2 are prevalent in breeding germplasm, FaRMp3 was identified in FVC 11-58, a reconstituted F. ×ananassa of diverse parentage that was previously identified as resistant to CR (Zurn et al. 2020). The resistance effect of FaRMp3 is stronger than FaRMp1 and FaRMp2 combined. The potential effects of pyramiding the three loci are currently undetermined. Moreover, FaRMp2 and FaRMp3 appear to reside in homoeologous regions that originate from two different ancestral diploids. Thus, marker-assisted pyramiding of FaRMp1, FaRMp2 and FaRMp3 may lead to stronger and more durable resistance against CR in new cultivars. Previous research in our group has shown that enhanced disease resistance can be effectively achieved through the combination of conventional and genomics-assisted breeding approaches. Developing subgenome- or allele-specific DNA markers in allo-octoploid strawberry is very challenging due to high levels of heterogenicity and complexity of homoeologous sequences in the subgenome. Our group recently developed a telomere-to-telomere (T2T) haplotype-phased reference genome of 'Florida Brilliance'. However, the high level of allelic and structural diversity in cultivated strawberry means that a single reference genome can only capture a fraction of relevant genes of interest. Without additional genomes, the search for candidate genes at all relevant loci will not be successful. In this project, new genomic tools and resources will be developed to enhance breeding for resistance to PhCR and CR.ObjectivesObjective 1. Characterizing Genomic Regions Conferring Resistance to Phytophthora Crown Rot and Charcoal Rot. Define genomic regions and fine-map QTL to discover subgenome-specific sequence variants underlying resistance to PhCR and CR.Objective 2. Developing Chromosome-Scale Haplotype-Phased Genomes Containing FaRPc2-H2 and FaRMp3. Generate high-quality haplotype-phased genome assemblies for elite breeding selections harboring loci conferring resistance to PhCR and CR.Objective 3. Designing Subgenome-Specific DNA Markers for Marker-Assisted Breeding. Develop high-throughput subgenome-specific DNA markers tightly linked to resistant alleles.?Objective 4. Accelerating Introgression of Multiple QTL Through Marker-Assisted Pyramiding. Combine multiple QTL for the resistance to PhCR and CR via marker-assisted pyramiding for increased resistance in elite breeding germplasm.