| The bioorganic fertilizer (BIO) containing Paenibacillus polymyxa SQR21, Bacillus subtillus SQR-9and Trichoderma harzianum SQR-T037has been shown to suppress Fusarium wilt disease, a severe soil borne disease, and to promote growth of watermelon and cucumber plants in greenhouse and field experiments. The present study was carried out to develop a mode of application of bioorganic fertilizer for improving the biocontrolling efficacy to Fusarium wilt. The interactions that occur between plant roots and the antagonistic microbe (P. polymyxa SQR21) were in detail studied to provide guidelines of how to use biocontrol agents more effectively in actual field situations. The results were obtained as follows:1. Nursery application of BIO could significantly increase the biomass of the watermelon plants compared with the treatments applied with organic fertilizer (OF) or with chemical fertilizer (CK). The fresh shoot weights of watermelon plant in BIO treatment were1.24and2.08times higher than both of OF and CK, and the dry root weights of watermelon plant in BIO treatment were1.22and1.67times higher than OF and CK, respectively; The length, surface area, volume, tips and average diam of roots were significantly increased by the application of BIO; The numbers of bacteria and actinomycetes were significantly increased by the application of BIO compared to the OF and CK, and the number of fungi was significantly decreased by application of BIO; Nursery application of BIO could effectively increase the photosynthetic capacity of watermelon leaves by increasing the content of photosynthetic pigments in watermelon leaves.2. Pot and field experiments were carried out to evaluate the efficacy of the BIO application on control of watermelon Fusarium wilt.1) The new mode of nursery application of BIO could significantly decrease the Fusarium wilt incidence, and the best biocontrol efficacy could be achieved by applying BIO in the nursery stage plus application in the transplanted soil.2) The way of BIO application could not only suppress the Fusarium disease, but also increase the biomass of watermelon and thus increase the yields. This could be attributed to the fact that nursery application of BIO could ensure the efficient colonization of the antagonist at the seedling stage and decrease the number of the pathogen in the rhizosphere after transplanting.3. The methods of color reaction in watermelon rhizosphere, GFP-labeling, petriplate cuture count were used to investigate the colonization-ability of SQR21in watermelon rhizosphere or on rhizoplane. SQR21could effectively colonize on the watermelon roots, propagate in rhizosphere as the growth of roots, and form biofilm on the surface of root tips of watermelon plants. These results indicated that SQR21could protect the watermelon roots from invasion by pathogenetic fungi, and could also support some basic theory for the research of mechanisms on biocontrol of Fusarium wilt by SQR21.4. A rhizobox experiment was carried out to investigate the distribution of F. oxysporum f.sp. niveum (FON) and its antagonistic bacterium (SQR21), in the soil at different distances away from the watermelon rhizoplane by real-time PCR. The results confirmed that BIO application significantly suppressed Fusarium wilt and promoted the growth of the watermelon plants. Application of BIO significantly enhanced the abundance of SQR21and reduced the population of FON in the soil near the rhizoplane as compared to the control. A maximal quantity (7.14lgcopies/g soil) of P. polymyxa SQR21was obtained in the rhizosphere under the BIO treatment, and this number was decreased with increasing distance away from the rhizoplane. The population of FON in the rhizosphere of the control was2orders of magnitude higher than under the BIO treatment, and with increasing distance away from the rhizoplane, the FON abundance increased in the SQR21inoculated treatments and decreased in the treatments without SQR21. In the soil far away from the rhizoplane (bulk soil), both FON and SQR21reached a level that was not statistically different between the treatments, which might be attributed to the background level of the infested soil. These results revealed that the BIO application with the developed method suppressed Fusarium wilt by the promotion of SQR21colonization in the rhizosphere, which largely reduced the population of the pathogen.5. Some organic acids were significantly detected in the root exudates of watermelon. Chemotaxis and swarming assays were performed to investigate the ability of these organic acids to induce the motility of P. polymyxa SQR21. The results showed that oxalic acid, malic acid and citric acid were present in the root exudates but only malic acid and citric acid could significantly induce motility in P. polymyxa SQR21. The maximal inducing ability was obtained from malic acid. Values for malic acid were3.9and1.5times higher than the control in the chemotaxis assay and the swarming assay, respectively. An in vitro experiment further confirmed that these organic acids could promote recruitment to P. polymyxa SQR21, thereby increasing the population in the rhizosphere.6. Conidial germination of FON decreased in the presence of root exudates from SQR21inoculated plants, while it was enhanced in the present of root exudates from FON inoculated plants as compared with control. Maximum germination was found in the root exudates from FON inoculated plant after30days, which was1.35times more than control. A split-root system was designed to verify the fact that the alterations of the exudation pattern in SQR21or FON inoculated watermelon roots were not only local, but also systemic. Eleven kinds of phenolic acids were detected in the root exudates by HPLC. The changes of cinnamic acid concentrations were positively related to the conidial germination. Assay of cinnamic acid on conidial germination of FON revealed that relatively high concentration of this acid stimulated the conidial germination, and vice versa. All of these results will help in the better understanding of the plant-microbe communication and will guide to improve the biocontrol strategies against Fusarium wilt of watermelon plants. |
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