| Meloidogyne incognita (Kofoid & White) is an important plant soil-borne pathogen which can infect hundreds of hosts, and it is difficult to control because of its high tolerance and ineradicable traits. No appropriate disease-resistant cultivar and chemical pesticides with low toxicity and high efficacy were used commonly. Meanwhile, using of biological control agents (BCAs) may provide a friendly alternative to chemical pesticides for plant soil-borne disease control. Therefore, it would be necessary to explore new biological control agents to control these plant pathgens.Twenty-six bacterial strains that had demonstrated antagonism to some fungal and bacterial pathogens were evaluated for their ability to inhibit the root-knot nematode. In the in vitro test, the 26 strains caused the J2 mortality rates of M. incognita varying from 1.3% to 77.8% and the inhibition rates of egg-hatching ranging from -16.5% to 87.4%. The 12 strains inducing J2 mortality over 40% were chosen for further greenhouse experiments and one of the remaining strains was randomly selected as a control, in which their biocontrol efficacy reached from 30.0% to 65.6%. On the other side, the 26 strains were tested for their protease and chitinase activities, which revealed that 20 of them had protease activities and 14 of them had chitinase activities. We investigated the relationships between the biocontrol efficacy and protease or chitinase activities. The biocontrol efficacy against M. incognita of the antagonistic strains were highly correlated to their in vitro protease activities (r=0.92), but not to their chitinase activities (r=0.06). We proposed that the in vitro protease activity of an antagonistic strain could be used as a parameter for selecting the BCAs against the root-knot disease. We further assessed the biocontrol efficacy of strains Bacillus sp. AR156 in field. Consequently, the biocontrol efficacy of AR156 reached 74.3%, in the field in Huai-an, Jiangsu, and 71.0% in Zao-zhuang, Shandong, respectively, which were significantly higher than that of abamectin:45.4% in Huai-an and 42.2% in Zao-zhuang. Biocontrol efficacy of Bacillus sp. AR156 with diferent application methods towards Meloidogyne root-knot were tested under field conditions. The application methods included (A) composting AR156 with rapeseed residue and made compost before application, (B) watering diluted AR156 into rhizosphere of plants, (C) spraying diluted AR156 into field, and (D) watering plants with dilution of AR156 preparation after plants treated by phoxime for 3 days. In field trials at Xuzhou in 2007, the total average control efficacy for treatments A, B, and D reached 74.4%,74.9% and 73.3%. And the yield increased 194.8%,193.1%, and 189.7%, respectively. However, method C, spraying, showed worse disease control efficacy than the other methods and the control efficacy was only 21.7% and the yield increased 39.7%. The average control efficacy of phoxime was 61.1% and the yield increased 155.2%.Strain AR156(Bacillus sp.) was isolated from Zhen Jiang forest soil in Nanjing, Jiangsu province. In order to investigate the mechanism of this biocontrol agent against plant disase, we studied its colonization and other activities. This strain could produce protease, cellulase, chitinase and siderophore. It also has the functions of phosphate-dissolving and nitrogen-fixing. Fourteen days after seed inoculation with 10 ml AR156 suspension with the concentration of 9.6×109 CFU/ml, the density of this strain in rhizosphere of tomato was 8.9×105 CFU/g soil. In addition, strain AR156 could induce the resistance of tomato against the diseases caused by Ralstonian solanacearum HB10. In green house experiments, on the 14th and 21th day after tomato plants were treated with strain AR156, the average biocontrol efficacy were 75.3% and 50.7%, respectively.Bacillus subtilis (Ehrenberg) Cohn have long been exploited for industrial and biotechnological applications. The completion of the sequencing and annotation of the B. subtilis 168 strain genome provieded a complete view of the Bacillus subtilis protein machinery. spoOA-deficient mutant of B. Subtilis 168 (mBs) contructed by using the pBGSCsp vector which could be integrated into spoOA locus within the B. subtilis 168 chromosome by single-crossover could not form biofilm. Meanwhile, another vector, pSGspOA, was integrated into amyE locus of the mutant (mBs) by double crossover, and we got the complementary strain named CmBs. PopW from Ralstonia solanacearum, is a member of the harpin group of proteins, eliciting hypersensitive cell death in non-host plants, inducing disease resistance in plants and enhancing plant growth. To expression and secreting PopW protein in Bacillus subtilis, we constructed recombinant expression vector pHTPOW, which carried p43 promoter, signal peptide element nprB, and the total PopW gen. The shuttle vector pHTPOW was transformed into competent cell of B. subtilis 168 and mBs separately, and two genetically modified strains, BsW and mBsW were constructed. The expression of PopW in two genetically modified strains was confirmed by SDS-PAGE and Western-blot analysis. In the in vitro test, on the 24 and 48 hous after treated with genetically modified strains, the J2 mortality rates of M. incognita were 53.4%, 79.6%,59.0%, and 89.7%, separately. On the 42th day, the biocontrol efficacy of strains (BsW and mBsW) reached 60.0% and 54.3%, respectively.In this study, bacteria GJ23 was used to control root-knot nematodes Meloidogyne spp., and it achieved a good efficacy. Previously, strain GJ23 showed great mortality of J2, in vitro. In the greenhouse tests, GJ23 could effectively inhibit gall formation in the roots of tomato. Under field conditions, the average biocontrol efficacy investigated on the 90th day after cucumber plants were treated with strain GJ23 was 63.3%, and yield increasement was 39.4%. As for control, the biocontrol efficacy and yield increasement of 1.8% abamectin were 35.4% and 13.2%, seperately. |
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