Isolation, Identification And Culture Condition Optimization Of The Strain Antagonist To Aspergillus Flavus And Preliminary Study On The Activity Of Supernatant Liquid

Fungus Aspergillus flavus is generally spread by producing spores, infection of plants and plant products, a variety of food and feed. Greatest harm is to produce aflatoxin aflatoxin. Aflatoxin (AFT) on the main staple food often appear in the Aspergillus flavus and Aspergillus Parasiticus with activity resulting from a toxic secondary metabolites. Because of the highly toxic to people,livestock and severe liver, the aflatoxin is the first of toxicity. Because of the great harm, the prevention of food safety control is particularly meaningful.Nowadays,the main research directions is using plant extracts and secondary metabolites of microorganisms to obtain toxic properties of antimicrobial substances or degradation. This study aimed to achieve antagonist metabolites biological control of aflatoxin is extremely toxic effects, and optimize the culture conditions for it and related properties of the metabolites of a preliminary study. The results were described as below:(1) One bacteria which have abvious antagonistic effect on Aspergillus flavus have been screened from the soil. The secondary metabolite of bacterium could inhibit the growth of Aspergillus flavus effectively and the rate of inhibitory is 37.88%. With regard to morphological features, physiological and biochemical test, and 16 S rDNA sequences analysis, the strain was identified. The results showed that the strain is a Bacillus atrophaeus. (2) Central composite rotatable design(CCRD) was used to optimize the culture conditions of a Bacillus atrophaeus inhibiting the growth of Aspergillus flavus.The effects of peptone, lactose, Mn2+, L-Cysteine monohydrochloride, pH, liquid medium volume, inoculum size, temperature, rotation speed and time were evaluated by using Plackett-Burman design. Results showed that the peptone and L-cysteine monohydrochloride as well as pH were the main affecting factors while the other factors had no significant effect on the production of fungistaticsubstance. Then, the central composite design and response surface analysis were used to determine the optimal levels of the main factors. The optimized conditions were as follows:peptone concentration 2.15 %, L-cysteine hydrochloride concentration 0.055g/L, pH7.75, lactin concentration 2%, inoculumconcentration 2%, liquid volume 30mL and the fermentation was processed at 30℃,190r/min for 90h. The rate of bacteriostasis was 56.30%, very closely matching the predicted value 55.71%. (3) This preliminary study of the metabolic products of B. atrophaeus protein concentration, the ability of inhibition to the growth of Aspergillus flavus compared with potassium sorbet and metabolites of Bacillus atrophaeus tolerance to temperature, pH, protease biological characteristics. The result showed that it has fungistasis on many different kinds of pathogenic fungi such as Fusarium graminearum and has broad-spectrum antibacterial activity also. 1mL fermentingliquor contained 0.5694mg protein, and the antifungal material contained in 1mL fermentingliquor as same as 0.013g potassium sorbet. Its antifungal activity was insusceptible after treatment under diferent temperature and pH. The active compounds could be degradated by papain partly. (4) The poisoned inhibition effect of coarse extraction products is studied in corn, peanut, soybean, rice on growth of Aspergillus flavus and yield of aflatoxin. Different kinds of grains were added 2,3,4mL fermentingliquor or bacterial suspension(1×107CFU) and 100μL spore suspension. After 5days, the concentration of positive groups were below 5ppb.