Pathogenicity Research Of Fusarium Fujikuroi With Screening And Functional Study Of Related BZIP Transcription Factors

Rice is one of the most important food crops in the world and more than 50% of the world population depends rice as their staple food.Rice bakanae disease what caused by Fusarium fujikuroi was one of seed born diseases.It caused wide distribution in rice growing area in the world.Bakanae disease caused excessive growth of seedlings,dead seedlings,dead plants,substantial yield reduction,huge economic losses,and even threatening food security.To understand the pathogencity of F.fujikuroi which come from different rice growth areas in China,such as Heilongjiang,Liaoning,Jiangsu,Zhejiang,Fujian provinces and Shanghai city,the invasion regularities of pathogens,especially the interaction between pathogens and hosts and their related biological functions,applied and basic relevant research were carried out,and the results were showed as follows:1.Differentiation of pathogenicity of bakanae disease pathogens F.fujikuroi194 strains of F.fujikuroi were obtained through isolated from the diseased samples collecting from Heilongjiang,Liaoning,Jiangsu,Zhejiang,Fujian provinces and Shanghai city.The invasion regularities,pathogenicity,and differentiation of 194 F.fujikuroi strains were investigated by artificial inoculation.Results revealed that the pathogenicity of most F.fujikuroi were moderate.The pathogenicity of these strains from different provinces and city were differentiated.The pathogenicity of F.fujikuroi from Zhejiang province were generally strongest,and significantly stronger than the pathogens from other provinces and city.The strains from Fujian Province were the weakest.There was no significant difference in the pathogenicity of F.fujikuroi from different regions in Zhejiang province.Differences in geographical environment,temperature,humidity and something like these might affect the pathogenicity of F.fujikuroi,whether from different provinces or city in China,or different regions in Zhejiang province.According to the pathogenicity of 194 fungi,20 strains with strong pathogenicity were selected,and 90%of them were from Hangzhou city,Zhejiang province.The strongest pathogenicity of F.fujikuroi was from Shaoxing city,Zhejiang province,named FF43(F.fujikuroi 43).2.Infection and colonization of F.fujikuroi in rice seedlingsFF43 strain was marked with enhanced green fluorescent protein(EGFP)gene.Through artificial inoculation,the infection and colonization of FF43 in the host was investigated by laser scanning confocal microscope.The results showed that F.fujikuroi could not invade the rice seedlings leaf sheaths,and the infection was mainly completed through the roots of rice seedlings.It meant that bakanae disease mainly caused by F.fujikuroi infecting the rice seedlings roots.In the process of infection,the hyphae penetrated into the cell tissue and continuously developed along the direction of the root tip.The infection was completed within 14 days.3.Gene expression and gibberellin synthesis of F.fujikuroiGibberellin is an important secondary metabolite which is synthesized by F.fujikuroi,playing an important role in the pathogenic process.Paclobutrazol regulates gibberellin synthesis in fungi,and exogenous gibberellin regulates the synthesis of gibberellin in F.fujikuroi through feedback repression.In this study,it was found that paclobutrazol significantly inhibited the vegetative growth and pigment synthesis of F.fujikuroi,and gibberellin reduced pigment synthesis of F.fujikuroi.On this basis,our study analyzed the effects of these two regulators on the gene expression of F.fujikuroi by transcriptome sequencing.The results indicated that exogenous gibberellin and paclobutrazol played an essential role in the gibberellin synthesis pathway-terpenoid skeleton biosynthesis pathway in F.fujikuroi by regulating gene expression what related to the mevalonate pathway.Exogenous gibberellin and paclobutrazol also affected gene expression of basic leucine zipper(b ZIP)transcription factors(TFs).The differences in the expression of b ZIP TF genes might affect the pathogenicity and gibberellin synthesis of F.fujikuroi.4.Genome analysis of the b ZIP transcription factor family of F.fujikuroiThe whole genome of F.fujikuroi was searched for gene sequences with b ZIP domain,and 29 F.fujikuroi b ZIP transcription factors were obtained.The intron,domain structure and phylogenetic relationship of b ZIP TFs were analyzed by bioinformatics,and it indicated that different domains might affect various biological functions of F.fujikuroi.By phylogenetic analysis and comparison with other eight fungi,it was found that the b ZIP TFs of F.fujikuroi were highly homologous with the eight fungi,and they might perform similar functions in F.fujikuroi.5.Functional analysis of the b ZIP transcription factor family of F.fujikuroiF.fujikuroi b ZIP transcription factor family contained multiple different domains.11 representative genes were selected out for further investigation.Through gene knockout,11 deletion mutants were obtained.Analysis of the biological characteristics and pathogenicity of the deletion mutants showed that the Ffb ZIP genes were involved in the vegetative growth,oxidative stress,osmotic stress,cell wall selective pressure,cellulose utilization,cell epidermal penetration and pathogenicity of F.fujikuroi.