Research On The Mechanism In Endospore Formation Of Paenibacillus Polymyxa And Its Preparation Of Microbial Pesticide

Paneibacillus polymaxa CF05 is a beneficial bacterium with the function of biocontrol of plant diseases and plant growth promotion.P.polymyxa CF05 produces endospores,which facilitate this bacterium formation of pesiticide.In this research,we focused on the formation of P.polymyxa CF05 endospores,and investigated the morphological and physiological characterisitics during the sporulation.Moreover,transcriptome analysis of P.polymyxa CF05 during sporulation was proceeded to discover the regulatory mechanisms.The CMC/PVAs spore formation was prepared for determining the stability of P.polymyxa CF05 and was tested for biocontrol of Carya cathayensis stem canker disease.The main results were listed as following.1.By using scanning electron microscope(SEM)and transmission electron microscope(TEM),the sporulation stages were observed.Three distinct stages,including vegetable cell,premature endospore and endospore,were determined according the cell morphological characters.During the vegetable cell stage,the cells were rod with 2.4×0.5?m,with flagellar.Cells were multiplied with binary fission.In the premature stage,the cells protruded at the ends,and the center of the bacteria began to circle,the surface of the bacteria began to be no longer smooth,and the size of the bacteria was about 1.5×0.8 ?m,At the endospore stage,the l cells were short rods,the ends were flat,and the surface of the bacteria had no flagellum and no ciliates.The size is about 1.0×0.9 ?m.2.RNA-seq was used to analyze the transcriptome profiles of P.polymyxa CF05 cells from vegetable cell stage and premature endospore stage.Results showed that the transcriptional levels of total 2039 genes were significantly different between vegetable cell stage and premature endospore stage.Among them,1069 genes were increased during the sporulation,and 970 genes were decreased.Noteworthy,there are 34 genes specifically expressed in the premature endospore stage and 2 genes specifically expressed in the vegetable stage.These genes are mostly involved in sporulation,carbon metabolism,cell division and membrane protein.It is precluded that those genes will be induced during the sporulation to contribute to the morphological formation,dormancy,and accumulation of response proteins for germination.3.We tested the effects of different carbon sources,nitrogen sources and potassium on the endospores production using single-factor experiment method.P.polymyxa CF05 cultured in the optimized medium(10g/L potato powder,20g/L glucose,5g/L beef extract,1g/L Mg SO4·7H2O,1g/L(NH4)2SO4,4.41g/L Ca Cl2·2H2O,0.6g/L KH2PO4,p H=7.0)produced the endospores with the concentration of 8.83×108CFU/m L in a short time.The fermentation showed the significant antagonism against plant pathogen Fusarium letersporum,Arthrinium phaeospermum,and Botrytis cinerea,which indicated that the optimized medium does not change the antibiotic activity of P.polymyxa CF05.4.Formulation of the strain endospores in carboxymethyl cellulose/ poly vinyl alcohol(CMC/PVA)blend exhibits a high stability over one year period.A two year field experiment shows that the CMC/PVA formulated strain reduces significantly the occurrences of both new cankers and developed cankers.P.polymyxa strain CF05 CMC/PVA formulation could be a potent environment friendly agent for the preventive biocontrol of C.cathayensis trunk canker.In conclusion,the morphological characteristics of the endospores of P.polymyxa CF05 were determined and the fermentation medium was optimized.The transcriptome profile of strain CF05 during the sporulation was declared.The results represented here would contribute to the development of strain CF05 as a potential micropesticide.