INTEGRATED GENOME-WIDE APPROACH FOR THE RAPID IDENTIFICATION OF NOVEL VERTICILLIUM RESISTANCE IN TOMATOES

Our long-term goal is to identify new genetic determinants of Verticillium Wilt (VW) resistance in tomatoes and enable the development of VW resistant cultivars. Verticillium wilt has long been a major limitation to tomato production, but with the loss of methyl bromide (MeBr) and the appearance of non-race 1 (Vdn) isolates of Verticillium for which there is no cultivar resistance, the need to identify and introgress genetic resistance is pressing. We have developed a team that over the past few years has put us in a unique and strong position to make significant advances in VW management. We have made significant progress in developing Verticillium resistance in horticulturally advanced NCSU tomato breeding lines as well as in the identification of the genetic variations associated with the resistance utilizing an integrated genome-wide approach. The prevalence of Vdn strains of Verticillium in the primary tomato production region in NC, with a consistent 100% infection rate on susceptible crops, has enabled us to evaluate and select for Vdn resistance under natural disease pressure through long-standing collaborations with tomato growers in on-farm trials. In the field, VW symptoms typically become more evident late in the growing season, and thus disease screening to evaluate VW resistance, particularly polygenic horizontal resistance conferred by multiple moderate resistant genes, is more appropriate from field screening rather than artificial inoculation of young seedlings in greenhouse experiments. Through extensive genetic analysis, we have shown that Vdn is likely controlled by a number of genes/QTLs. In other work, we have shown that non-race 1 isolates of Verticillium can be further classified into race 2 and race 3 using the tomato rootstock hybrids resistance to race 2 isolates ('Aibou', 'Ganbarune-Karis', and 'Bowman'). We refer to VW isolates that are pathogenic as race 3 on these lines and race 2 if they are non-pathogenic. We have developed molecular markers to identify different strains of the pathogen and showed that race 3 isolates are the most predominant strains of Verticillium in the primary tomato production region in NC. This might be the reason why race 2 resistant tomato rootstocks fail to provide protection against Verticillium in western NC. Preliminary greenhouse evaluations have shown that polygenic resistance presence in NC tomato germplasm might be effective against both race 2 and 3 isolates and we will refer to it as non-race 1 (Vdn) resistance hereafter. From a practical perspective, the incorporation of resistance to non-race 1 strains of VW will broaden the disease-resistant spectrum of tomato cultivars and consequently will minimize economic risk for growers in NC, the U.S., and worldwide.The following objectives express a two-tiered approach proposed for this research:1) identify and fine map non-race 1 resistant loci in three NC tomato breeding lines2) Utilize VW (non-race 1) resistance to develop new tomato hybrids stacked with additional disease resistancesWe will also utilize an innovative combination of the most recent genetics and genomics approaches to achieve our objectives. We will utilize a mapping-by-sequencing approach followed by map-based gene cloning employing residual heterozygous line to clone novel Vdn resistant gene(s). Instead of generating low-resolution recombination maps by PCR markers, the Allegro targeted-genotyping by sequencing approach will be used to construct a high-resolution recombination map of the NILs to increase map-based cloning accuracy for the first time in this proposal.Therefore, this proposal responds directly to the research needs recognized by growers, extension agents, and researchers; furthermore, it directly addresses AFRI program goals in most of the Farm Bill priority areas. We aim to address the needs recognized by AFRI as "Plant health and production and plant products" program area priority by focusing on the latest plant breeding, genetics, and genomics research to ensure that U.S. agriculture is prepared to meet the grand challenges facing the world. Our innovative approaches of identifying and utilizing host resistance in tomato production are key to producing more food with less impact on the environment. We have built a team and tomato breeding platform, enabled by genome-wide and innovative breeding schemes, data analysis, and knowledge of molecular and biological processes, for quick transfer of traits into elite tomato cultivars. In collaborations with small and commercial tomato growers, as well as private seed companies, multiple on-farm trials extended our tomato breeding efforts into public and private domains and enabled us to train for current and future growers, plant breeders, and researchers.