| Rice-duck Farming Ecosystem (RDFE) is effective in controlling weeds and pests, reducing methane emission in paddy, and has advantages of zero-tillage, enhancing rice yield and quality. It is reported that RDFE is effectual in controlling Rhizoctonia solani(R.solani). The research was divided into two aspects to explore mechanism of RDFE restricting R.solani. On one hand, the influence of duck movements on rice and R. solani was researched, on the other hand, the effects of extractions and microbes from duck beak, excrement, feather on rice and R.solani were also studied. Both researches were aimed at exploring the factors and mechanisms of RDFE controlling R.solani and pursueing new approaches in R.solani controlling.1 Research on effective factors and mechanisms1.1 Effects of RDFE on the number of R.solani sclerotiumThe number of R.solani sclerotium was investigated in both Duck-raised and non-duck-raised paddies where initial number of R.solani sclerotium were equal. During late tillering stage and full heading stage of rice, the sclerotium number were 357,000 pieces/hm2 and 419,000pieces/hm2 in Duck-raised paddies which were 48.97%and 43.46%lower than non-duck-raised paddies with the number of 729,000 pieces/hm2 and 964,000pieces/hm2. This proved that the reduction number of pathogenic bacteria and the disease controlling were contributed by duck movements.1.2 Effect of closing mud on R.solaniIn order to learn the effects of closing mud on R.solani, the research was carried out by closing mud on the surface of basal leaf sheath of rice. Mud was closed manually on the surface of basal leaf sheath and then R.solani was inoculated, the morbidity was between 3.08～10.61%, disease index was 5.42-11.82 after the treatment, which was significantly lower than Validamycin treatment (3.88～55.36%, 5.66～18.40) and blank control (3.97～86.22%,19.94～60.72). And the control efficiency was 72.80～81.21%. Peroxidase activities were significant higher than blank control but lower than Validamycin treatment in the treatment of mud closing then inoculation, which also lowered soluble sugar content but there was no significant difference among the treatments.1.3 Effects of extractions from duck beak, excrement, feather on chitinase activityPrevious research has showed that there was positive correlation between disease resistance of rice and its chitinase activity. Compounds that were applied on rice were extracted from duck beak, excrement and feather by ethylacetale and carbinol, and then chitinase activity of rice was tested. Chitinase activity got to the maximum of 124.7 U·g-1·FW-1·min-1 on the 8th day after treated by compound extracted from duck excrement by ethylacetale, significantly higher than blank control(77.11 U·g-1·FW-1·min-1) (p<0.05, p<0.01); Chitinase activity got to the maximum of 62.9 U·g-1·FW-1·min-1 on the 8th day after treated by compound extracted from duck excrement by carbinol, which had no significant difference with 62.9U·g-1·FW-1·min-1 of blank control (p>0.05, p>0.01); This proved that the enhancement of chitinase activity was contributed by the compound extracted from duck excrement by ethylacetale, presumably that the containment of certain somatomedin and compounds could trigger rice disease resistance. While, there were no significance found from the same operations of duck feather and beak.1.4 Effects of microbes separated from duck feather, beak and excrement on chitinase activity323 strains of bacteria and 126 strains of fungi were separated from duck feather, beak and excrement by dilution plate method, bacteria B106 and fungi F123 were from duck feather, B75 and fungi F66 were from duck beak, bacteria B117 and fungi F16 were from duck excrement, whose suspensions were treated on rice respectively. Chitinase activity got to maximum of 42.24U·g-1·FW-1·min-1,42.23 U·g-1·FW-1·min-1 at the 3rd and 7th day after B117 treatment, significantly higher than CK 41.74U·g-1·FW-1·min-1,41.69 U·g-1·FW-1·min-1(p<0.05,p<0.01). This proved that microbes from duck excrement could enhance chitinase activity, while others not.1.5 Effects of extraction from duck feather, beak and excrement on R.solani Control efficiencies of extractions from duck beak, feather and excrement by ethylacetale and ethanol were tested on R.solani via plate antagonism method, the inhibition effect of which were 13%,8%,6%,7%,88.64%,84.11%respectively; EC50 of excrement-ethanol treatment in 24h was 26.11mg/ml; Crude extraction of duck excrement was separated into petroleum-extraction and ethylacetale-extraction, and the ethylacetale-extraction got relatively higher control efficiency,89.0%, with EC50 of 15.52mg/ml, and the petroleum-extraction got relatively lower control efficiency; Ethylacetale-extraction were filtered and separated by means of silica gel column chromatography and 5 components were acquired, and the L2 fraction had significant control efficiency, with 89.58%and 80.36%in 24h and 48h, which was identified as phenolic acid and mercaptan via GC-MS. This proved that compounds existed in duck secretion could control R.solani in RDFE.1.6 Effect of microbes from duck feather, beak and excrement on R.solaniAntagonisms of B106, B75, B117, F123, F66 and F16 against R.solani were tested via plate-confront method. No antagonism were found in strains of B106, B75, F123, F66 and F16, while B117 isolated from duck excrement showed high antagonism against R.solani, and A168 strain was acquired via further screening of B117. This proved that antagonistic bacteria could be isolated from duck secretion in RDFE.2. Research on screening, identification, biological property and antagonism of A1682.1 Research on screening, identification and biological property of A168By purifying B117 for 100 times via plate-antagonistic method, A168 strain was selected, identified and biologically characterized. A168 was rhabditiform, Gram-positive, central spore, peritrichate. It can utilize Glucose, Arabinose, Xylose and mannitol to produce acid, and is negative in Glucose gas producing test, Amylohydrolysis test, anaerobic growth test, Tyr degradation test, Phenylalanine deaminase test, while positive in Hydrogen nitrate reduction test, Casein degradation test, Catelase test and VP reaction test. It had homology of 99% with its consanguinity Bacillus according to 16S rDNA analysis.16S rDNA sequence was registered in GenBank with the accession number of GQ118339. Population of A168 had maximum in 60h cultivation, its optimum temperature was 28℃, and it lived well at pH7.0. Sucrose, Glucose, Mannitol, a-lactose, D-xylose, maltose, D-galactose and D-fructose could be utilized as carbon source, and tryptone, peptone, beef extract, yeast extract, casein and salmiac could be utilized as nitrogen source by A168.2.2 Research on process of active materials extraction and fungistatic mechanism of A168Optimum extraction process were worked out:ratio of ethylacetate and zymotic fluid was 4:1, temperature 28℃,36h constantly ultrasound extraction. EC50 and EC90 of extracted active material against R.solani were 2.15mg/mL and 237.86mg/mL; mycelium twisted after active material treatment; Inhibition ratio of soluble protein had maximum of 24.45% and germination ratio of sclerotium had minimum of 80.36% at treatment concentration of 35mg/mL; Pathogenicity of R.solani decreased along with increasing concentration of active material, and inhibition ratio reached to 80.29% at concentration of 45mg/ml; Effect of active material on electrical conductivity of mycelium was not found yet. This proved that active material of A168 could control R.solani by inhibiting mycelium growth, protein synthesis and sclerotium germination, as well as reducing pathogenicity.2.3 Research on fermentation condition and optimum substrate for A168Optimum fermentation and cultural condition were researched via orthogonal test:bean-cake powder 2%, glucose 7%, CaCO3 0.4%, NaCl 0.35%, peptone 6%. Optimum in vibrating flask:48h,28℃, cell age 24h, inoculation volume 6%, rotate speed 200r/min, liquid capacity 210ml (500ml in vibrating flask), pH7.0. Optimum in lOL-fermentor:VC 180L/h, rotate speed 200r/min,28℃, nascent pH6.5, duration 72h.2.4 Research on colonizing regulation and control efficiency of A168 on riceColonizing regulation and field control efficiency of A168 were explored via strain recycling marked method. Friction was optimum for colonizing; A168 had largest population at tillering stage in rice after inoculation, then full headin stage and mature period had smallest; Basal sheath had denser population than root and stem; Live strain could be obtained after 32 days of inoculation; Field control efficiency reached to 78.47%, significant higher than Validamycin treatment.This proved that A168 had a higher colonizing capacity and control efficiency on rice.2.5 Research on growth promotion, disease resistance and yield improvement effects of A168After treated by A168 suspensions, root length, plant height, leaf length and fresh weight of rice seedling were respectively increased by 12.35～30.95%,26.52～30.77%,16.37～20.02%and 10.26～29.52%, compared to blank control; A168 had better growth promotion effects in rice variety "Lemont" and "Luliangyou996" than in "ChuangfengNo.1"; Compared to blank control, "Lemont" had higher content of IAA, ZR4, GA3 and lower content of ABA, and had higher activity of PAL, POD but no significantly different activity of CAT and APX after treatment of A68 suspensions; A168 suspensions increased rice yield, with increasing rate of 13.13% on "Luliangyou 996", and then "Lemont" and "Chuangfeng No.1". These proved that A168 had effect of promoting growth, improving disease resistance and yield on rice.2.6 Research on screening, purification and identification of active materials in A168Chemical reaction, Ammonium sulfate precipitation, DEAE-SepharoseFF and FPLC Phenyl FF lyophobic chromatographic column, HPLC C18-purification were used in order to study, isolate, purificate and identificate the active materials in A168. Protein and peptide were isolated from filtrate of A168, and such active material was named L-158, and then contributed to further separation and purification. Via SDS-PAGE electrophoresis analysis,15KD of L-518 was found; Via mass spectrum analysis, L-518 was identified to have charge-to-mass ration of 15006.899. Sample L-518 was not well resistant to high temperatures, and stable when exposed to protease K while instable to trypsase and pepsase. Considered these characters were similar to polypeptide, L-518 was presumably identified to be polypeptide.
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