Improving key root traits in sugar beet: Fusarium resistance

The challenge of the twenty-first century is to produce enough food to meet population demands without extending land or damaging the environment. Combining a maximum number of desirable traits such disease resistance, greater yield, and high quality is a desirable goal for plant breeders. The development of resistant crop genotypes is essential to ensure global food security, make the plant more useful and avoid crop losses. The development of molecular markers linked to the target traits is needed to predict phenotypic variation based on genotype. Marker-Assisted Selection (MAS) can reduce costs and the time required to obtain new cultivars by comparing selection only based on phenotypic evaluation. Single nucleotide polymorphisms (SNPs) are widely used as genetic marker. Sugar beet (Beta vulgaris L.) is the second source of world sugar supply and is grown in all temperate zones. The crop is attacked by many pathogens and among these, the soil-borne fungus Fusarium oxysporum causes severe sugar beet damages. Two different formae speciales have been reported in sugar beet, F. oxysporum f. sp. betae that causes Fusarium yellows, and F. oxysporum f. sp. radicis-betae that causes Fusarium root rot. Disease symptoms are characterized by wilt and yellow leaves that normally die as the disease progresses. Internal symptoms consist of a brown or grey brown vascular discoloration and in the case of root rot, there is a back external rot in the primary root. Sugar beet varieties are susceptible to F. oxysporum, which can cause a lower root yield and reduce sugar quality. No genetic studies have been done up to now, so no genes or quantitative trait loci (QTLs) conferring resistance to F. oxysporum in sugar beet have been reported. The aims of this work were (i) to investigate the response of a wide collection of sugar beet lines to F. oxysporum f. sp. betae, (ii) to identify resistant lines suitable for future breeding efforts and (iii) to discover molecular markers linked to the Fusarium resistance that could be considered for use in marker-assisted selection (MAS) programs. The first part of the thesis is a literature review of sugar beet breeding achievements, including the discovery of monogermity and cytoplasmic-genetic male sterility (CMS) that allowed the release of hybrid varieties. The review also focused on the breeding progresses against diseases obtained with classical and molecular methods using sources of resistance from wild beets. Next-generation sequencing (NGS) technologies with the recent release of the full sugar beet genome sequence are also reported. Incorporation of genomics into conventional sugar beet breeding programs is essential to obtain important yield achievements in sugar beet. The second part was aimed at screening a wide range of sugar beet lines to identify the different effect to F. oxysporum f. sp. betae inoculation and to select resistant and susceptible lines. To achieve this, 29 sugar beet lines were screened under greenhouse conditions with two highly virulent isolates belonging to different genetic sub-groups. The third part regards an experiment conducted to evaluate the response of different sugar beet breeding germplasm to isolates of F. oxysporum f. sp. betae. In the previously tested lines, an unusual root rot was observed, normally reported in cases of infection with F. oxysporum f. sp. radicis-betae. Eight susceptible lines, from USDA-ARS (US) and UNIPD (University of Padova, Italy), were inoculated with three different isolates of F. oxysporum f. sp. betae, the causal agent of Fusarium yellows. All inoculated lines developed disease symptoms, but severe root rot was observed only in the susceptible UNIPD lines inoculated with isolates that had never caused root rot in the USDA germplasm. In this work, an unusual root rot was reported for the first time that seems to be caused not only by the isolates, but is also due to a germplasm effect. The fourth part was aimed to identify molecular SNP markers linked to the Fusarium resistance in sugar beet. A candidate gene approach was used on susceptible and resistant lines to achieve this goal. Five resistant gene analogues were screened by means of a high-resolution melting (HRM) analysis and two allelic variants, within two genes, were significantly associated to Fusarium resistance. Sanger sequencing allowed the discovery of two SNP markers linked to the resistance. These two SNPs were significantly associated with the resistance and were mapped on the exon of Bv7_171470_ojty and Bv2_043450_zhxk, respectively