Research On Epidemical Mechanism Of Rice-Duck Integrated System Suppressing Rice Sheath Blight And Application Potential Of Antagonistic Bacteria SU8

Researches in this paper were conducted aiming to investigate the epidemical mechanism of Rice-Duck Integrated System (RD) suppressing Rice Sheath Blight. Experiment was carried out between two treatments, rice combined with ducks (RD) and conventional rice field without ducks rearing (CK) in early season rice paddy, to investigate the variations of sclerotia in floodwater and on rice plant, microclimate10cm above the waterline in rice paddy and activity of protective enzymes in rice plants. The results showed that the floating sclerotia in floodwater in RD was86%～91%lower than that in CK, and adhering sclerotia in rice plant in RD was67%～78%lower than that in CK. The relative humidity tested significantly lower and light intensity tested significantly higher in RD. The temperature in the early rice growth stages in RD was slightly lower than that in CK, but it was significantly higher (32.3～36.5℃) in the middle stage rice growth stages. The polyphenoloxidase (PPO) activity in RD were lower than that in CK, but the enhanced activity of phenylalanine ammonia-lyase (PAL), peroxidase (POD) and Chitinase was observed in different stages of rice growth in RD, especially the Chitinase which showed higher activity in all investigating days. At dough stage of growing rice, the disease index in RD was76%lower than that in CK, and the rice yield in RD was40%higher than that in CK.During the above researches, author isolated anagonistic bacteria strain SU8from rhizoshperic soil in RD, which exhibited superior antifungal activity against Rhizoctonia solani. In the fallowing research, bacteria stain SU8was studied based on morphological, physiological and biochemical characteristics, sequence of16S rDNA genes and phylogenetic analysis. And, in order to ascertain its functioning mechanism in Rhizoctonia solani control, the active compound from the sterilized fermentation liquid was isolated and purified. The molecular structure of the compound was detected through GC-MS,’H-NMR(300MHz) and13C-NMR(300MHz). The results show that:1. According to the result of morphological research, strain SU8was shaped as long rod, the length ranged1～1.5μm, G negative. The feature in petri dish and its physiobiochemcial characters matched the description of standard Pseudomonas aeruginosa. The family tree based on the16S rDNA sequence showed SU8had99%similarity with other known strains of Pseudomonas aeruginosa, and the sequence was recorded in GeneBank, obtained sequence number HQ283487.2. According to the test of antibacterial spectrum, strain SU8had inhibitory effect on Fusarium graminearum, Phytophthora parasitica, Colletotrichum capsici, Magnaporthe oryzae, Ustilaginoidea virens, Rhizoctonia solani, Alternaria alternata. Its sterile zymotic liquid was used to evaluate suppressive effect on pathogenicity of two diseases, and pot culture experiments were also conducted to evaluate its control efficiency. And the control efficiencies both reached up to70%on infected rice plants in pot culture. Strain SU8showed superior potential as a bio-control agent in suppressing ShB and RB.3. Identification of the activity compound in the metabolites of Pseudomonas aeruginosa SU8is the main target of this research. Through the selection of crude extraction method, solvent extraction was selected to be the most effective method by ethyl acetate to sterile zymotic liquid with ratio of1:1,3repetitive operation. Two times of Silica column chromatography were established to obtain the purified Sample-4, with the first elution agent system of petroleum ether:chloroform as ratios of10:1、10:2、10:3、10:4、10:5、10:8、10:10、8:10、5:10,and second elution agent system of petroleum ether:chloroform as ratios of petroleum ether:ethyl acetate as ratios of10:2、10:4、10:6、10:8、10:10、0:1.Through bio-test, Sample7-4was selected out to be active compound, which weighed21.5mg and was light-yellow crystal. HPLC test showed that the purity reached up to97%.4. Sample7-4was analyzed by GC-MS. According to the database, the molecular formula was given as C13H9N3O. Through the’H-NMR(300MHz) and C-NMR(300MHz) detection, the active compound of Pseudomonas aeruginosa SU8was identified to be Phenazine-1-carboxamide, PCN and the molecular structure:, and acquired spectrum parameters showed highly resemblance with the report by R. Sunish Kumar.According to the given results,①the density of the sclerotia was considerably decreased, contributing the inhibited proliferation of primary source of infection, which led to the reduction of diseased plant rate and diseased hill rate. In the rice canopy, reduced relative humid, illumination intensity and dynamics of temperature created the condition adverse to the growth of Rhizoctonia solani. And, under the condition of duck intervention, the rice plant cells underwent the physiobiochemical changes which enabled the cells to generate the induced resistance to plant disease, like the enhanced activity of PPO, PLA, POD and Chitinase. Those changes made positive contribution to the suppression of rich sheath blight.②Pseudomonas aeruginosa SU8has broad-spectrum antifungal activity. The control efficiency on rice sheath blight in culture pot indicates strain SU8has the valuable property as an biological control agent, which provides the quality to be used for the development of live bacteria agent. The isolation and purification of the active compound from sterilized fermentation liquid help to give referable information for active compound extraction. And, the identified molecular structure of active compound contributes to ascertaining the antifungal mechanism of strain SU8, which also provides the important information for develop stable, safe and efficient bionic fungicides for rice sheath blight control.