| Wheat is a cereal crop which is widely planted all over the world.Fusarium head blight caused by Fusarium graminearum is one of the main diseases of wheat,not only leads to the decrease of wheat yield,but also accumulates toxin in the grain,such as Deoxynivalenol(DON)and Nivalenol(NIV),which seriously affects the quality of wheat and makes people and animals acute poisoning,causing huge losses to the safety production of wheat.At present,the preventive and control measures against wheat scab include chemical control and breed disease-resistant varieties,but long-term use of chemical agents will enhance the resistance of Fusarium graminearum and cause environmental pollution.Due to the lack of wheat cultivars resistant to Fusarium Head Blight and the complex inheritance of resistance to Fusarium Head Blight,safe and effective control measures are urgently needed.Biological control has gradually become a hot topic for scholars because it is non-toxic to crops and friendly to environment.The antagonistic characteristics of microorganisms have a good prospect for the control of Fusarium Head Blight,which is worth further study.The use of Bacillus to control Fusarium graminearum is safe and stable,while the mechanism of interaction between Fusarium graminearum and Bacillus is still unclear.According to this topic,the following studies have been carried out:256 bacterial strains were isolated and purified from wheat ear samples,and 78 antagonistic bacteria were screened by plate confrontation method.Among them,strain YB-130 showed significant antagonistic effect against Fusarium graminearum,and was identified as Bacillus velezensis by morphological and molecular biology.The whole genome sequencing of strain YB-130 showed that the genome was composed of a circular chromosome without plasmid.The total length of the chromosome was3980767 bp,and the content of G+C was 46.46%.It was predicted that there were 4105 CDS,and the total length of the coding gene was 3551001 bp,covering 89.20% of the genome.It contained 86 t RNA and 27 r RNA,and a total of 12 gene clusters encoding secondary metabolite synthesis were predicted,of which 4 gene clusters had unknown function.Compared with Fusarium graminearum p H-1 cultured alone,the inhibition rate of spore germination was 67.58%,the inhibition rate of mycelial growth was 71.8%,the content of DON was decreased by 69.27%,and the relative expression levels of DON biosynthetic genes Tri3,Tri5 and Tri8 were down-regulated by 6.5,10 and 2 times respectively.The results of bacteriostatic spectrum showed that the inhibition effect on Alternaria solani and Bipolaris sorokiniana(Sacc.)was good,and the inhibition rates were 72.71% and 68.22%,respectively.Transcriptome sequencing results of Fusarium graminearum interaction with strain YB-130 showed that Fusarium graminearum p H-1 obtained 22.56 G Clean reads,and the Q20 and Q30 values were all above 98% and 94%,and the sequencing error rate of base position was all lower than 1%.A total of 6439 differentially expressed genes were obtained,among which 3169 genes were significantly up-regulated and3270 genes were significantly down-regulated.Among GO functions,there were 37,22 and 13 annotations to molecular function,biological process and cell composition,respectively.Among them,the biological process had the highest annotation abundance,while the cell composition had the lowest annotation abundance.KEGG enrichment analysis showed that the genes were significantly up-regulated in 11 processes,including ribosome,aminoyl t RNA biosynthesis,RNA transport,amino acid biosynthesis and RNA polymerase,gene down-regulation was mainly concentrated in9 processes,including proteasome,peroxisome,steroid biology,terpenoid main-chain biosynthesis,endocytosis and fatty acid metabolism.In this study,the fatty acid synthetase FGSG＿11656 gene was screened by using the interaction transcriptome data of Fusarium graminearum p H-1 and strain YB-130.Then,the FGSG＿11656 gene knockout box was constructed by Spilt-marker method,and the protoplast transformation was mediated by PEG.The deletions mutants were screened by a certain concentration of ampicillin,hygromycin resistant plate and PCR positive and negative primers.The phenotypic observation and pathogenicity determination of the obtained mutant showed that compared with the wild-type PH-1,the pigment yield of the ΔFGSG＿11656 mutant decreased,the mycelium margin became thinner,and the growth rate of the mutant was not significantly different.Compared with wild-type p H-1,the spores of ΔFGSG＿11656 deletion mutant were partly transparent at the apex,and the size of the spores was not significantly different.The spores production and dry weight of mycelia were reduced by 54.54% and 22.77%,respectively.The DON content of wild-type strain was up to 115.2±2.85 μg/m L.The yield of DON in ΔFGSG＿11656 deficient mutant was only 57.13±2.39 μg/m L,and the DON content was decreased by 50.67%.The coleophotile infection test showed that the incidence of wild-type PH-1 was as high as 91.67%,and the incidence ofΔFGSG＿11656 deletion mutants was only 53.33%.These data can provide scientific basis for further study on the interaction mechanism of Fusarium graminearum responsive strain YB-130. |
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