Study On Pathogenicity Appreciation Of Sesame Fusarium Wilt Pathogens And Sesame Genetic Resistance To SFW

Sesame(Sesamum indicum L.,2n=26), which belongs to Sesamum genus in Pedaliaceae family, is one of the most important oilseed crops. Sesame is infected by Sesame Fusarium wilt (SFW) annually with about15%, as well as more than30%in serious seasons, which gives significant impact on sesame seed yield and quality. SFW is caused by Fusarium oxysporum f. sp. sesame (FOS) and present in sesame in any growth stages. To set up the appreciation technique of pathogenic level of FOS pathogens and disclose mechanisms of FOS pathogeneis and sesame resistance on FOS, we collected and isolated257FOS strains from infected sesame plants in China; the appreciation technique of pathogenic level was determined by indexes screening; the pathogenic level of20strains of FOS were determined. Furthermore, we had built the genetic transformation system of FOS by Agrobacteria tumefaciens mediated transformation method (ATMT), in which hygromycin resistance gene and green fluorescent protein (GFP) gene were transformed into genomes of FOS. We had observed the infection process using a FOS strain transformed with GFP gene. Moreover, we investigated genetic resistance on FOS with six generations of a sesame population (Zhongyou1134×Sichuan Rongxian black sesame) in FOS artificial nursery. Main results are shown below.1. We established the pathogenicity appreciation technique of FOS, after omparing and analyzing effects of inoculation concentration of strains, investigation stage and disease symptom types on sesame seedlings. Results showed that the appropriate concentration of FOS stain suspension was about1×106spores per milliliter solution for soil inoculation. Before sowed, aseptic sesame seeds were preferred to germinate. The appropriate survey period was20-22days after inoculation, as the disease symptom presented almost in1week and disease indexes would varied evidently for2-3weeks before unchanged. We surveyed twenty strains of FOS using the above technique and screened four virulent strains and3weak pathogneic strains. Pathogenic levels of different strains differred from each other, as well as one strain showed different pathogenicity among different cultivars.2. We set up and optimized the genetic transforming system of FOS pathogen using Agrobacteria tumefaciens mediated T-DNA insertion technique (ATMT). Genes encoding for green fluorescent protein (GFP) and Hygromycin were successfully transferred into genomes of FOS. The highest transformation efficiency of100-200transformants per106spores were obtained, as the fusarium oxysporum conidia suspension of1×106spores per milliliter were mixed with the Agrobacterium bacteria with OD6oo=0.6and cultured for24hours under the conditions of120rpm shaking and22℃, and co-cultureed for48hours in IM medium with75μmol/LAS. We had built the first mutant library of FOS with982mutants in China using the above technique. We analyzed the phenotype and morphology of116transformants. Results showed that main mutated traits included colony morphology, growth rate, colony color, pigment color, sporulation capacity, spore morphology and pathogenicity. We found two weaker pathogenic strains and one stronger pathogenic strain. These research results give the basis for further genes determination related with pathogenicity.3. To explore the interaction relationship between sesame and Fusarium oxysporum f. sp. Sesame, a nonsense mutant strain of FOS,12G-3are selected for infection observation on sesame seedlings. Results indicated that the infection happened in12hours. In the infection process, Fusarium oxysporum firstly attached to the intercellular spaces of root epidermal cells, then infected the surrounding tissues and invaded into lateral tissues of root hair zone and the main roots of sesame seedlings at last. A large number of hyphae were observed colonized at the infected spots of roots and woven into meshes.4. We inoculated the isolated and purified strains of FOS, HSFO08005、HSFO07003、 HSFO08030、HSFO09067, into the artificial nurseries in sesame SFW resistance research. Infected percentage of the six generations (P1、P2、F1、BC1、BC2and F2) of the population of Zhongyou1134×Sichuan Rongxian black sesame was investigated in strains-mixed nursery (2010, PingYu) and sole strain nursery (2011, Yuan Yang). Resistance inheritance on FOS of the population was analyzed using the major gene+polygene model and analysis method. Results suggested that①Resistance of on SFW was controlled by two major genes with additive effects plus polygenes with additive dominance effects in mixed nursery.②Resistance of the above population was controlled by one major gene with additive effects plus polygenes with additive dominance effects in single strain nursery. Inheritance of Resistance in Sesame Fusarium Wilt was complex and greatly impacted by environment. In a conclusion, the above results formed the foundation for future researches on pathogenecity of FOS and sesame resistance on SFW and disease-resistance breeding.