| Fusarium head blight(FHB)caused by Fusarium graminearum complex is an important crop fungal disease.F.graminearum infects wheat,which not only causes serious yield reduction of wheat,but also pollutes wheat and its products with mycotoxins such as deoxynivalenol(DON toxin)secreted by F.graminearum,threatening human and animal health.In recent years,the occurrence of FHB showed a trend of moving westward and expanding northward,the epidemic risk increased,and the prevention and control faced continuous pressure.Due to the lack of wheat varieties resistant to scab,chemical control has always been the main means of scab control.However,long-term single use of chemical pesticides will not only bring environmental pollution,but also cause drug resistance of field strains.Mining green and efficient biocontrol strains,as an effective supplement to chemical control,is the only way to upgrade diversified control strategies.Endophytic bacteria are a kind of bacteria colonized in plants for a long time,which may have multiple functions such as promoting growth and resisting diseases,and are important microbial resources for biological control of plant diseases.In this study,an endophytic bacterium CR7 with antagonistic effect on F.graminearum was isolated from plant roots and identified as Burkholderia gladioli by 16 S sequencing,and the growth inhibition rate of F.graminearum was 44.4%.CR7 treatment did not affect the conidia germination,ascomycete and ascospore formation of F.graminearum,but the spore production of F.graminearum decreased seriously.Through the pathogenicity test,we found that strain CR7 could significantly inhibit the pathogenicity of F.graminearum.In view of the fact that CR7 has great effects on the growth,sporulation and pathogenicity of F.graminearum,we carried out further studies on this strain to reveal its mechanism.It was found that the metabolites of CR7 did not affect the conidia germination of F.graminearum,but could significantly inhibit the mycelial growth and pathogenicity of F.graminearum.Under the treatment of metabolites shaken with YEPD,the mycelium growth of F.graminearum was inhibited by 57.8%,and the pathogenicity was reduced by 73%.It is speculated that the strain plays a role in biocontrol through its metabolites.Furthermore,we determined the stability of metabolites under acid-base,heat,ultraviolet and protease treatment.The results showed that the metabolites were relatively stable in acidic environment,while the bacteriostatic effect decreased in alkaline environment;after UV irradiation for 1hour,the bacteriostatic rate of metabolites did not change compared with the control group,but decreased with the increase of irradiation time;the inhibitory effect of metabolites treated with high temperature and protease K on F.graminearum decreased by nearly half compared with the control.Furthermore,the genome of strain CR7 was sequenced and assembled,and the gene clusters of secondary metabolites synthesis of genes on the genome were predicted on the anti SMASH website,and a total of 21 gene clusters were predicted.According to the biological characteristics of strain CR7,we selected the gene cluster lactimidomycin for knockout,but the inhibitory effect of knockout mutant on F.graminearum was no different from that of wild type.We also used transposon insertion to identify the genes with antibacterial function in strain CR7.After successfully constructing the mutant library,we screened 18 mutants with reduced growth inhibition rate of F.graminearum through bacteriostatic experiments.Sequencing showed that the defects of antibacterial function of these mutants may be related to coding genes such as GTP binding protein,LSU ribosomal protein,SSU ribosomal protein,50 S ribosomal protein,ATP-dependent protease and arginine succinate synthase.In order to explore the application potential of CR7,we tested the colonization effect of strain CR7 after root irrigation treatment of wheat.It was found that strain CR7 could colonize in wheat tissue,indicating that it had the most basic colonization ability.We further tested whether CR7 could induce resistance to wheat.The results of wheat panicle inoculation experiment showed that CR7 treatment could not induce disease resistance in wheat.In addition,In addition,we tested the bacteriostatic effect of CR7 on other pathogens of wheat diseases(Cochliobolus sativus 、 Colletotrichum asianum 、 F.pseudograminearum 、Gaeumannomyces graminis var.tritici 、 Rhizoctonia cereadis and F.asiaticum).The experimental results showed that strain CR7 and its metabolites had good bacteriostatic effects on these six pathogenic fungi,and the best inhibitory effects were G.graminis var.tritici and F.pseudograminearum,with inhibition rates of 56.3% and 60.3% respectively.To sum up,B.gladioli CR7 and its metabolites have good antibacterial effect on F.graminearum and other pathogenic fungi,and the strain can colonize wheat tissues,which has certain application potential. |
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