| Mulberry bacterial wilt is a worldwide soil-borne plant disease caused by Ralstonia solanacearum.Mulberry bacterial wilt is widely distributed in mulberry fields all over China,among which mulberry fields in southern China are the main affected areas,which has seriously restricted the sustainable development of sericulture industry.At present,the control of mulberry bacterial wilt mainly adopts chemical control and agricultural control.The longterm application of chemical agents will make pathogenic bacteria resistant,increase the difficulty of control and cause serious damage to the ecological environment,which is not a long-term effective control measure.Biological control has gradually become a new strategy for plant disease control because of its advantages of green environmental protection,safety and efficiency.Therefore,more and more attention has been paid to the screening of antagonists,the exploration of antibacterial mechanism and the separation and purification of active substances.In this study,an actinomycete strain ZM-1 with significant antagonistic effect on R.solanacearum was isolated and screened from the rhizosphere soil of healthy plants in the diseased mulberry garden.In order to further explore the control effect of antagonistic actinomycetes on mulberry bacterial wilt and its control mechanism,the microbial classification and identification,optimization of liquid fermentation conditions,stability analysis of fermentation bacteriostatic substances,analysis of biocontrol characteristics and pot experiments were carried out.The main results are as follows :1.Using R.solanacearum as the indicator bacteria,the strain ZM-1 with significant antagonistic effect on R.solanacearum was isolated and screened from the rhizosphere soil of healthy plants in the diseased mulberry garden by gradient dilution separation method and agar diffusion method.The diameter of the inhibition zone was 17.5 mm.The antibacterial spectrum of strain ZM-1 was determined by plate confrontation method.It was found that it had significant antagonistic effects on Aspergillus flavus,Rhizoctonia solani,Fusarium pseudograminearum,Ceratocystis fimbriata,Staphylococcus and Fusarium oxysporum.2.By observing the morphological characteristics and cultural characteristics of the antagonistic strain ZM-1,combined with the determination of physiological and biochemical characteristics and molecular biological identification,it was finally determined that the strain was Streptomyces hachijoensis,the aerial hyphae were soft,the basal hyphae were not diaphragmatic and not broken,the spore filaments were spirally arranged,and the spores were oval.3.The liquid fermentation conditions of antagonistic strain ZM-1 were optimized by single factor test and response surface method.The optimum fermentation medium was determined as follows : glucose 35.5 g / L,yeast extract 12.5 g / L,tryptone 10 g / L;the optimum fermentation conditions were as follows : fermentation time 120 h,inoculation amount 4 %,temperature 30 °C,p H = 8.1,shaking speed 200 r / min.The antibacterial circle diameter of the optimized sterile fermentation filtrate reached 21.298 mm,which significantly improved the antibacterial activity of strain ZM-1 against R.solanacearum.4.Using R.solanacearum as the indicator bacteria,the stability of antibacterial substances in the fermentation filtrate of strain ZM-1 was determined.The results showed that the antibacterial substances in the fermentation filtrate of the strain had good thermal stability and good stability under ultraviolet irradiation,but the acid-base tolerance was poor,which had certain reference value for the separation and purification of antibacterial substances in the later stage.5.In this study,the biocontrol characteristics of antagonistic bacteria ZM-1 were analyzed by using hydrolytic enzyme qualitative detection plate.The results showed that strain ZM-1 could produce fungal cell wall hydrolytic enzymes-lipase,protease,cellulase,and also produce siderophore,which had the characteristics of promoting plant growth.6.The pot experiment explored the preventive effect of different concentrations of fermentation filtrate of antagonistic bacteria ZM-1 on mulberry bacterial wilt and the growthpromoting effect on mulberry seedlings.The results showed that the fermentation filtrate of antagonistic bacteria ZM-1 diluted 40 times had the best preventive effect on mulberry seedlings,and the incidence rate was reduced by 53.3 %.At the same time,when the fermentation filtrate of antagonistic bacteria ZM-1 was diluted 10 times,the mulberry seedlings were irrigated,which had a growth-promoting effect on mulberry seedlings. |
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