Potential And Analysis Of The Antifungal Compounds Produced By Bacillus Sp. B21 As A Biopesticide To Control Rice Blast

Rice blast,a devastating fungal disease,causes tremendous loss of rice production and quality worldwide.Up to 30%of the rice crop is destroyed by rice blast each year,causing a concerning situation for food production.To control rice blast,chemical pesticides are usually employed in rice production.However,the extensive use of pesticides puts the ecological balance and human health under great threat,meanwhile,Magnaporthe oryzae often can develope resistance to fungicides.Biological control,which is considered an alternative method for disease control,is a new method that uses the antibacterial activity of organisms or their bioactive metabolites to control varieties of diseases.The biological control of rice blast has the advantages of low cost and environmental friendliness when compared with the application of fungicides.In this study,both antifungal activity and biocontrol efficacy of Bacillus sp.were studied.The results were as follows:1.One of the endophytic bacteria,which were isolated from Osmanthus fragrans Lour.,showed obviously antifungal activity against M.oryzae.The strain was named B21and identified as Bacillus safensis by using morphological,physiological,biochemical and molecular biological methods.The experiments indicated that B21 had a strong adaptability to the environment and simple requirement for nutrition.Twenty four hours post inoculation,the strongest antifungal activity could be detected.2.The culture filtrate showed clear antifungal activity against M.oryzae in a dose-dependent manner.In particular,Methanol extract from culture filtrate inhibited M.oryzae growth with an IC₅₀ of 16.56?g/ml,which was significantly(P<0.01)lower than that of carbendazim with an IC₅₀ of 25.16?g/ml.According to the physical and chemical experiments,the culture filtrate was stable at a wide range of pH(1-9)and temperatures(40-100?).3.Three antifungal compounds were isolated from culture filtrate by using organic extraction and chromatographic column separation.The NMR data indicated that these three compounds belonged to iturin and surfactin lipopeptide families.The exact mass was further confirmed by HPLC-MS and the molecular weights were 1042.5,1070.5 and1035.7,respectively.From the above,the antifungal compounds were identified as iturin A2 with the molecular formula C₄₈H₇₄N₁₂O₁₄,iturin A6 with the molecular formula C₅₀H₇₈N₁₂O₁₄4 and surfactin with the molecular formula C₅₃H₉₃N₇O₁₃.4.Both iturin A2 and iturin A6 are lipopeptides,which have a strong inhibitory effect on filamentous fungi.After treated with iturin(iturin A2 or iturin A6),the hyphae of M.oryzae showed the inhibition of apical growth accompanied by morphological abnormalities with swollen and vacuolated tips that can be stained easily by PI.These results indicated that iturin A2 and iturin A6 inhibited the growth of hyphae by changing the membrane permeability.In addition,iturins could also inhibite spore germination and cause collapse in appressoria.5.Evaluation of the biocontrol efficacy against rice blast was conducted on detached leaves and in field trails.The results indicated that leaves treated with fermentation broth or culture filtrate had smaller disease spots than controls.Furthermore,B21 showed better biocontrol efficacy than carbendazim in field trails.All results suggested that strain B21 and its bioactive compounds had their advantages and could be developed into a biopesticide for the biocontrol of rice blast.