Screening And Inhibitory Effect Of Bacillus Amyloliquefaciens FB-16against CROP Phytophthora SP.

Plant fungal disease is the main reason for the declining of yields and quality ofagricultural products. In its prevention and treatment process, the large number of long-termuse of chemical pesticides has led to serious environmental and food safety issues. Therefore,looking for the non-pollution measures to prevent and control the plant diseases, screeningand developing new microbial fungicides for plant fungal diseases is very important. Thisresearch aimed at isolating and screening the microbes with strong antagonism against cropPhytophthora from the soil samples of healthy tobacco rhizosphere in high incidence tobaccofields, and investigation on its taxonomic status, antifungal activity and antifungal mechanismto get a new microbial pesticide resources against Phytophthora.From the30soil samples of healthy tobacco rhizosphere in high incidence tobacco fieldsin Shaanxi Province, such as Longxian, Jingyang, etc.,347strains of bacteria were isolated byplant dilute. Antagonist of the bacteria was screened with several fungal pathogens, such asPhytophthora nicotianae、Phytophthora capsici Leonian、Botrytis cirerea、Physalosporapiricola、Fusarium oxysporum f. sp. vasinfectum、Pyricularia oryzae by plant pathogens. Theresultss showed that only eight bacterium had strong antifungal activity, and strains F16aswell as F205, had better antifungal activity. After rescreening the above two strains by filterpaper, we found that the F16has better antifungal activity, and the inhibition zone diametersagainst Phytophthora nicotianae and Phytophthora capsici Leonian were29.7mm and27.0mm.The antagonistic bacteria FB-16was identified based on morphological, physiologicaland biochemical characteristics, and16S rDNA phylogenetic analysis. The resultss showedthat the morphological characteristics, physiological and biochemical characteristics of FB-16was consistent with the pattern bacterial of Bacillus, which could resist acid, alkali, salt, andhigh temperature, and its sequence is similar to Bacillus amyloliquefaciens in the NCBI, ofwhich similarity is99.728%. Based on the above characteristics, the strain FB-16wasidentified as B. amyloliquefaciens.The inhibition rate of mycelium growth and the spore germination were determinedusing the methods of mycelium growth rate and the spore germination test. The results showed that the inhibition rate of mycelium growth of FB-16fermented liquid againstPhytophthora nicotianae was95.02%, and the inhibition rate of its spore germination was94.60%. The antifungal mechanism experiments showed that the FB-16could bring about themycelium tumescent, abnormal, branching increases, and plasma overflow. The bonsaiexperiments in greenhouse indicated that the control effect of active product of FB-16againsttobacco black shank was up to69.96%, which had potential value in biological control againsttobacco black shank.Besides, some primarily study had been carried out on the antagonism of FB-16strain.We found that FB-16has protease and amylase activity, speculating that the protease oramylase which it secreted played a role in the antifungal. The active metabolite of sterilefermentation of FB-16had good stability against heat, strong resistance against UV, andstrong adaptability to acid-base environment. The antifungal metabolite by stain FB-16precipitated by25%ammonium sulfate indicated the highest effective inhibition in theextraction process of active substances.