| Potatoes are solanaceae of herbaceous perennial tuber plant, and China is one of the world’s largest potato production bases. Potato used widely with high economic value. Potato now becomes the fourth-largest food crops after rice, wheat, and maize on the world. The potato late blight caused by Phytophthora infestans (Mont.) de Bary is one of the most serious diseases which can lead to steams and leaves death and steam tuber rot. At present, the control measures of potato late blight are mainly chemical control, but the water, air and soil are polluted by chemical pesticide seriously and through the food chain could enter human body, to endanger public health. Using biological control method is expected to avoid the above shortcomings, and has broad prospects.The actinomycete strain NB-8, which was isolated from the vegetable soil of NingBo in our lab, and its living and fermented liquid had a strong antagonistic effects on mycelial growth of P. infestans according to the research of our lab. The inhibitory rate of living and fermented liquid reached 92% and 80%, respectively. And NB-8 strain was identified as Streptomyces albus. The active component exsited in NB-8 fermentation could be dertermined as a kind of non alkalic antibiotics. However, the inhibition of NB-8 strain on P. infestans dropped significantly in later experiments, even decreased to 55%, although the cause was not clear. Therefore, using a single factor experiment and the orthogonal experiment, optimization of medium and fermentation conditions for NB-8 strain was investigated in this experiment. The antimicrobial activity of NB-8 strain and fermentation was also studied. The activities of some resistant-related enzymes in restrained P. infestans of different inhibition area were measured. The results were as follows:1.The fermentation conditions of NB-8 strain were optimized. The results showed that optimal fermentation conditions of this strain were seed liquid cultured in Gause 1 medium for 5d with amount of 1.5% and a volume of 125mL/250mL, the initial pH7.0, 50 r/min, 34℃, 12h dark/12h light, cultured 10d. Under the optimal fermentation conditions, the inhibitory rate of fermented liquid on P. infestans reached 81%, which increased by 20% in comparison with pre-optimization.2. Based on Gause’s No 1 synthetic medium, optimization of medium for NB-8 strain was studied. The results showed that the optimal carbon, nitrogen, and NaCl concentration for single factor were yeast creams, lactose, and 10g/L, respectively. And the optimal fermentation medium for multi-factors were affirmed by orthogonal tests. The best medium combination was that lactose 10g/L, yeast creams 4g/L, K2HPO4 0.5 g/L, MgSO4 1.0 g/L. On this basis the inhibitory rate reached 89% .3 . The optimal concentration and stability of NB-8 fermentation liquid against P.infestans was understood. The antifungal activity enhanced with increasing the fermentation liquid concentration during the range of 5.3-84.8 mg/mL. The lowest inhibitory rate was 70.21% with dilution 2 times (5.3 mg/mL), the highest was 95.78% with enrichment eight times (84.8 mg/mL). The inhibitory effect overall declined as the storage time prolonging, but the inhibitory rate was still above 75% after stored 12 months. And the fermentation liquid had good stability for most salt ions, such as MgSO4,MnSO4,K2HPO4,(NH4)2SO4, the inhibitory rate was still above 64% treated with ZnCl2 which had the strongest influence to the fermentation liquid.4. The antimicrobial spectrum of NB-8 strain living and fermentation liquid were cleared. NB-8 strain living had different effect to the growth of 22 tested fungi. The inhibitory rates were more than 50% for 11 fungi of total, especially for Alternaria alternata, A.brassica, and YH-1, the inhibitory rates were up to 70%. But the inhibition of fermentation liquid for most of the tested fungi was poor except for Botrytis cinerea. The obvious inhibiting effect of the fermentation liquid to the tested bacteria was observed except SR13, the inhibition zones were more than 12 mm. Among them, inhibiting effects to both Sporosarcina sp. and Bacillus subtilis were the strongest, the inhibition zones were up to 20 mm.5.The inhibitory mechanisms of NB-8 fermantation on P.infestans were preliminarily studied. The results showed that the NB-8 fermentation liquid could result in deformation of P. infestans mycelium. It was found that mycelia uneven thickness, coarse, cell inclusions gathered in P. infestans which was inhibited by NB-8 fermentation liquid. The NB-8 fermentation liquid against P.infestans could not keep inhibitory activity for 7 days. The activities of some resistant-related enzymes in P.infestans mycelia of restoring growth, which were restrained in different inhibition area (0, 10, and 20 mm from the edge of inhibitory belt) were measured, respectively. The activities of SOD, CAT, PAL and the content of MDA, as well as soluble protein in restrained fungus decreased evidently, especial MDA and SOD, dropped by 60-90% and 53-85%, respectively. The content of soluble protein reduced by 55-64%, and the activities of PAL and CAT declined by 48-56% and 38-64%, respectively. It is contrary to plant whose related enzyme activities usually increase after infection or intimidation. |
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