Research On The Mechanism In Biological Control Of Paenibacillus Polymyxa And Optimization Of Fermentation Condition

Paenibacillus polymyxa,a endospore-forming bacterial,is a plant growth-promoting rhizobacterium.The P.polymyxa CF05 was isolated from an ancient tree Cryptomeria fortunei located in Tianmu Mountain and has a potential commercially useful biocontrol agent.For lying a foundation of further definitions and exploitation of biopesticide,this study clear bio-control and growth-promoting mechanism,grope for fermentation process and preparate of microbial pestcide of P.polymyxa CF05.The main results were listed as following.1.Dual-culture inhibition and double layer culture methods were performed for measuring antagonistic effect against 8 phytopathogenic fungi(Botrytis cinerea,Fusarium moniliforme,Fusarium oxysporum f.sp.vasinfectum,Ceratobasidium cornigerum,Fusarium oxysporum,Alternariasolani solanisorauer,Cercospora circumscissa,Thanatephorus cucumeris and four phytopathogenic bacteria(Agrobacterium tumefaciens,Pseudomonas.marginalis,Erwinia.carotovora subsp.carotovora,Curtobacterium flaccumfaciens).The results demonstrated that P.polymyxa CF05 exhibited highly antagonistic effect against phytopathogenic fungi and bacteria.2.Growth-promoting effect of P.polymyxa CF05 on tomato and control effect of P.polymyxa CF05 on tomato root-knot disease and Fusarium wilt were tested by greenhouse trials.The results showed that control efficiencies of root gall and fusarium wilt were 64.4% and 78.24%,respectively.Meanwhile,treatment of P.polymyxa CF05 had remarkable impact on tomato growth,fresh weight growth rate is 29.4% and dry weight growth rate is 38.4%.3.Growth-promoting effect of P.polymyxa CF05 on Dendrobium officinale Kimura et Migo and control effect of P.polymyxa CF05 on leaf spot disease were tested by greenhouse trials.The results showed that control efficiencies were 63.6%,fresh weight growth absolute rate is 74.6%,and increase on the number of lateral buds is 7.1 while the control is 2.2.4.Tests of root-colonizing ability of P.polymyxa CF05 with antibiotic resistance method,indicated that concentrations of the bacterium in tomato rhizosphere were 8.0×103CFU/g root after 25 days of inoculation of tomato seeds with the spores suspension at 108 CFU/mL.5.The culture supernatant of P.polymyxa CF05 were confirmed the presence of IAA by HPLC.The results showed that P.polymyxa CF05 produced IAA in tryptophan biosynthesis pathway and the highest concentration of IAA in supernatant is 5.35 ?g/ml.6.Enzyme activities of PAL,PPO,PODand phenol content were determined in tomato roots with different treats.Experimental results revealed that P.polymyxa CF05 activated PAL?PPO?POD?SOD in tomato roots and maintained for 3-4 days.It is indicated that P.polymyxa CF05 induce plant systemic resistance.7.Pesticide-sensitive test of P.polymyxa CF05 was performed and the results indicated that strain CF05 exhibited resistance to abamectin,but have high sensitive with some bactericide,such as tebuconazol.8.The complete genome map of P.polymyxa CF05 was achieved.The result shows that P.polymyxa CF05 genome,G+C% is 45.47%,contains a single circular chromosome of 5 762 698 bp and no plasmid.The chromosome codes 5 032 ORFs that were predicted with Glimmer3.9.By multi-factor orthogonal experiment and single-factor test,the optimum medium and its fermentation condition of P.polymyxa CF05 was studied.Results showed that optimum fermentation medium component for strain P.polymyxa CF05 was 20.0g/L corn flour,10.0g/L bean flour,10.0g/L fishmeal,5.0g/L K2HPO4,0.5g/L CaCl2,0.5g/L MgSO4.And the optimized fermentation condition was described as: inoculation rate 2%,pH 7.0,temperature30?.At last,the optimized technology was applied 100 L fermentation tun production,optimized technology allowed the yield of P.polymyxa CF05 to 5×1010 CFU/ml.In conclusion,this study illustrated that P.polymyxa CF05 have significantly antagonistic effect against phytopathogenic fungi and bacteria and control effect on tomato root-knot nematode disease,Fusarium wilt and leaf spot disease of D.officinale Kimura et Migo.Meanwhile,P.polymyxa CF05 had remarkable impact on growth of tomato and D.officinale Kimura et Migo.This study clear bio-control and growth-promoting mechanism of P.polymyxa,including efficient root colonization,IAA production,as well as inducing systemic resistance.Development of fermentation procedure for P.polymyxa CF05 laid the foundation of development of P.polymyxa.