Screening And Identification Of NF011 Against Fusarium Graminearum And Its Biological Control Mechanism

Fusarium Head Bheat（FHB）caused by Fusarium graminearum（Fg）is a serious hazard.It not only causes a large area of production reduction,but also produces Deoxynivalanol（DON）,etc.,Seriously affect the yield and quality of wheat.When the intake of DON exceeds1000μg/kg,it will poison humans and animals and cause symptoms of vomiting.At present,due to the single field management method,the breeding of resistant varieties requires a long period,and chemical agents have also caused serious harm to human health and the environment.Therefore,screening high-efficiency Fg antagonistic bacteria and preparing new,high-efficiency and stable biological control agents are of great significance for effectively reducing wheat head blight,reducing toxin hazards and ensuring the safety of humans and animals.In Jingmen,Hubei,we found that wheat in the rice-wheat rotation area and the wheat in the jade-wheat rotation area had different conditions of head blight,and then the disease index statistics were carried out during the flowering period of the wheat.We found that the disease index and DON content of wheat in the jade-wheat rotation area were significantly lower than those in the rice-wheat rotation area.The main bacterial communities at the level of rhizosphere soil microbes are all Pseudomonas,and the number of Pseudomonas in the soil of the jade-wheat rotation area is significantly higher than that in the soil of the rice-wheat rotation area.In an experimental field in Zhongxiang City,Hubei Province,we collected wheat and root soil samples in rice-wheat rotation and jade-wheat rotation fields.After separation and screening,a total of 33 bacteria were found to have antagonistic effects on Fg.Among them,the samples were selected from the soil of the jade-wheat rotation.NF011 has the best antagonistic effect on Fg,with an inhibitory rate of 64.52%.After morphological identification,physiological and biochemical characteristics,molecular identification and phylogenetic tree analysis of NF011,NF011 was identified as Pseudomonas aeruginosa.In order to study the antagonistic effect of NF011 bacterial solution against pathogenic fungi,NF011 was tested with 12 different plant pathogenic fungi.The results showed that NF011 has antagonistic effects on 12 plant pathogenic fungi,indicating that NF011 has a broad spectrum of antagonistic phytopathogenic fungi.ability.The conditions of NF011bacteria liquid against pathogenic fungi were optimized,the optimum p H was 6 and the optimum temperature was 28℃.The effect of NF011 on antagonizing the growth of Fg and the production of DON was studied.It was found that the protoplasts of Fg ruptured after treating the hyphae suspension of Fg with NF011 at a concentration of 10⁸ CFU/m L for 1 day.The silk shrinks,thereby inhibiting the growth of Fg hypha.NF011 at 10⁸ CFU/m L can also significantly inhibit the germination and growth of conidia of Fg.After applying NF011 bacterial liquid,it can obviously inhibit the growth of Fg in wheat ears and kernels,thereby inhibiting the content of DON in wheat kernels,which is similar to the effect of the chemical pesticide carbendazim.The 20%NF001 fermentation supernatant was studied to antagonize the activity of Fg,and it was found that 20%of the NF011 supernatant had an inhibitory effect on Fg,with an inhibition rate of 33.97%.In order to separate,purify and identify the small molecule substance that NF011antagonizes Fg,the medium under the co-culture of NF011 and Fg was cut,and then leached with ethyl acetate,extracted and separated with distilled water,and then used Silica gel column chromatography was used for three purifications,thin layer chromatography and HPLC were used to identify the purity,and relatively pure antibacterial small molecules were obtained.Finally,LC-MS and nuclear magnetic resonance were used for identification,and the analysis showed that the antibacterial small molecule was Phenazine-1-carboxamide（PCN）.The PCN produced by NF011 antagonizes the growth of Fg and found that the minimum inhibitory concentration of PCN is 32μg/m L,and the inhibitory rate reaches 66%.It was also found that 32μg/m L PCN can significantly inhibit the growth of Fg hyphae and the germination of spores,which is the same as NF011.The content of PCN produced by different Pseudomonas strains and the ability to antagonize Fg were studied.It was found that among the 9 Pseudomonas strains,only Pseudomonas aeruginosa NF011 had the extremely obvious ability to antagonize Fg,and only NF011 produced a large amount of PCN.,PCN content reached 80.24μg/m L.The conditions for NF011 to produce PCN were optimized.The results showed that the most suitable initial medium was KMB medium,the best time was 72h,the best carbon source was mannitol,the best nitrogen source beef extract and peanut meal,and the best temperature was 20℃,the optimum p H is 6,the best metal ion is Mn²⁺,the best amino acid is Arg,and the best added strain is Alcaligenes faecalis NF037.The field effect of NF011 on the growth of winter wheat and the control effect of scab was investigated.The results showed that field experiments showed that NF011 can effectively inhibit the growth of Fg and reduce the content of DON.