The Screening Of Biocontrol Agents Against Fusarium Head Blight And The Research On Frenolicin B Against The Disease

Wheat(Triticum aestivum L.)is one of the most important food crops.Fusarium head blight(FHB)of wheat caused by Fusarium is a worldwide fungal disease.In recent years,due to the changes in climate and planting methods,FHB has been widespread and outbreak posing a serious threat to global wheat production and food safety.Due to the lack of FHB-resistant seed resources,chemical control is still the main measure against FHB.However,the long-term heavy use of chemical agents has caused environmental pollution and endangered biological health,in addition it has caused severe drug resistance.Biological control is more friendly to the environment,not easy to produce resistance,and the resources are abundant.Recentiy,biological control by microbial resources and its metabolites has become a major trend under the concept of green agriculture.In this study,6 separate mediums were used to isolate actinobacteria from wheat habitats.The high-efficiency biocontrol isolates against FHB were screened in vitro and vivo assays.Determined the fermentation activity of biocontrol strains.In combination with the results of secondary metabolites predicted by whole genome sequences and the separation and purification of active compounds,identified the main active metabolites of biocontrol strain.Then the activities,control effects in fields and antagonistic mechanism of the main active conpound were further explored.The main results of this study are as follows:(1)827 atinobacterium were isolated from wheat samples collected from the wheat fields of Anhui and Henan provinces,of which Streptomyces accounted for the major part.Statistical analysis and comparison of the abundance and number of actinobacteria separated from different locations in different samples were carried out.It was found the actinobacteria diversity of rhizosphere soil,root and ear of wheat infected with FHB and the actinobacteria abundance of rhizosphere soil and ear of wheat infected with FHB were significantly higher than the healthy group,while the abundance of root was lower than the healthy group.Moreover,the actinobacteria diversity and abundance of rhizosphere soil were much larger than the endogenous actinobacteria of wheat,and the largest in diseased wheat rhizosphere soil,while the smallest in wheat ear.(2)10 specific atinobacterium isolated from the wheat habitats were identified as new species of genus Streptomyces(4),Jiangella(1),Microbsipora(3),Sphaerimonospora(1)and Pseudonocardia(1),respectively.Among them,6 new isolates were isolated from the rhizosphere soil of disease wheats with 4 Streptomyces sp.nov.,and biocontrol agent NEAU-H2 was one of them.(3)104 active strains against Fusarium graminearum PH-1 were selected in vitro assay,and the activities of 20 strains were greater than 50 %,which were selected for antifungal activity test in vivo.Based on the activity result of vitro and vivo assays,the morphological characteristics and phylogenetic position of 20 biocontrol isolates,strains NEAU-H3 and NEAU-H17 had strongest activities and distincted from other strains.(4)Biocontrol agent NEAU-H3 has a strong antagonistic effect against PH-1,in addition it suppressed a variety of plant pathogens and had cellulose and lignin degradation,IAA production and other activities.Moreover,the sterile fermentation supernatant of NEAU-H3 had a good prevention effect against FHB.Therefore,it was a potent biocontrol agent with potential to produce novel antifungal compounds.(5)In combination with the results of secondary metabolites predicted by whole genome sequences and the separation and purification of active compounds,the main active metabolite of strain NEAU-H3 was determined to be pyranaphthoquinone compound Frenolicin B.(6)Frenolicin B had strong inhibitory activities against FHB,and showed good control effects in the field tests.Furthermore,Frenolicin B had strong inhibitory effect s against variety of plant pathogenic fungi.According to the results of the interaction resistance test s,Frenolicin B had no interaction resistance with carbendazim which was commonly used in FHB control.Frenolicin B is a new prevention agent against FHB.(7)It was found that the mycelia of PH-1 were uneven in thickness,swelling and distortion,and the internal structure of the mycelia changed significantly with pyknosis cytoplasm and varying density after Frenolicin B treatment by electron microscopy.(8)In order to further explore the mechanism of antagonism,transcriptome assay of PH-1 was tested after Frenolicin B treatment.It was found 2,520 DEGs,and the genes related phosphatase,and energy metabolism significantly down-regulated after Frenolicin B treatment,while the genes associated with stress response and nucleic acid repair were significantly up-regulated.According to GSEA of the KEGG pathways and GO terms,Frenolicin B significantly inhibited a variety of PH-1 metabolic processes,among which nucleotide and energy metabolism were the main targets of Frenolicin B.(9)The active metabolites of strain NEAU-H17 were isolated and purified,and the main active metabolites were identified as analogues of azalomycin F: 2-Demethylazalomycins F4 a and 2-Demethylazalomycins F5 a.It has been widely used in plant pest control and medical research.In this study,the compound showed a strong inhibitory effect against F.graminearum,indicated that it has the potential of developing agricultural antibiotics to control FHB.