| Applying phosphate solubilizing bacteria (PSB) was one of the effective methods to utilize fixed P in soil. In order to keep stabile phosphate solubilizing activity after application, PSB should be rhizosphere efficient strains which were suitable for the rhizosphere of plant. In this study, PSB were screened from rhizosphere of elephantgrass and maize using modified Pikovskaya (PVK) plate method, and phosphate solubilizing ability (PSA) of these strains was determined by measuring the soluble P concentration in National Botanical Research Institute's Phosphate (NBRIP) broth. In addition, PSB were tested in the laboratory for their ability to colonize the rhizosphere of maize seedlings in sterile quartz sand so as to find rhizosphere efficient strain. Then a pot trial was carried out in greenhouse to evaluate whether the laboratory test can predict beneficial effects of PSB on maize. Finally, we studied mechanism of rhizosphere efficient strain X14 suitable for the rhizosphere of plant from the perspective of effect of maize root exudates on its rhizosphere colonization and PSA and effect of synthetic sugar mixtures (in analogy to root exudates of maize) on its PSA, and rhizosphere colonization was focused on swarming, chemotaxis and growth. The main results obtained were as follows:1. Twelve PSB were isolated from rhizosphere of elephantgrass and maize, and we selected three strains designated as X14, X32, and Y21 according to spore-staining results and PSA in liquid medium, whose soluble P concentration in NBRIP broth with Ca3(PO4)2 as insoluble P source were 123.72 ?g·mL-1,542.58 ?g·mL-1 and 618.77 ?g·mL-1, respectively. Based on morphological and biochemical characteristics and phylogenetic analysis of 16S rDNA, strain X14, X32, and Y21 were identified as Bacillus megaterium, Acinetobacter parvus and Pseudomonas psychrotolerans.2. The phosphate-solubilizing activity of strain X14 was significantly lower than strain X32 and Y21, but it was found to be a better rhizosphere colonizer than strain X32 and Y21, whose final population in the rhizosphere of maize seedlings expressed as CFU·g-1 was an order of magnitude higher than strain X32 after 8 days in equal inoculum size. Under greenhouse conditions, application of PSB remarkably improved plant diameter, shoot height, fresh and dry weight, and P uptake of maize as compared to the control, but there was no significant difference between treatments inoculated with PSB.3. We collected maize root exudates under gnotobiotic conditions. Maize root exudates could promote swarming and chemotatic response of strain X14. The organic acids in maize root exudates were detected by high performance liquid chromatography (HPLC), and effect of the identified organic acids on the chemotaxis of strain X14 was preformed. The results showed that there were six organic acids in maize root exudates, including oxalic acid, malic acid, acetic acid, succinic acid, trans-aconitic acid and propionic acid. It was found that four organic acids could significantly induce chemotatic response, and the maximal inducing ability was obtained from propionic acid, whose relative chemotaxis index was 3.9, followed by trans-aconitic acid, succinic acid and malic acid, respectively. Moreover, we studyed the growth and PSA of strain X14 in modified NBRIP broth A with maize root exudates as sole carbon sources. The results showed that strain X14 could grow with maize root exudates as carbon sources, its population reached the maximal densities of 2.14×106 CFU·mL-1 within 1 day, which was an order of magnitude higher than initial inoculum size, and remained stable after slight decrease. The soluble P concentration in modified NBRIP broth with maize root exudates as carbon sources of strain X14 was significantly higher than all the controls, which included control without carbon source and strain X14, control without carbon source but inoculated with strain X14 and control with root exudates of maize but uninoculated with strain X14.4. The growth and PSA of strain X14 in modified NBRIP broth B with synthetic sugar mixtures (in analogy to root exudates of maize) as carbon sources were tested. After 7 days incubation, the population of strain X14 was an order of magnitude higher than initial inoculum size, and the soluble P concentration in the strain X14 culture was 93.44 ?g·mL-1, which was 11.8 times as high as that in the control. HPLC analysis results showed that there were 45.23 mmol·L-1 lactic acid,19.00 mmol·L-1 malic acid,4.20 mmol·L-1 gluconic acid,0.41 mmol·L-citric acid and 4.77 ×10-3 mmol·L-1 fumaric acid in the culture of strain X14. In artificial P solubilization trail, there were significant differences among solubilization of Ca3(PO4)2 by strain X14, mixtures of organic acids and HC1, and the P concentration in broth of organic acid mixtures and HC1 were 77.07% and 13.10% of strain X14. Solubilization of Ca3(PO4)2 by these five organic acids were significantly different, and lactic acid, malic acid and citric acid played a dominant role. |
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