| Phosphorus is an essential nutrient for crop growth and agricultural production mainly uses phosphorus fertilizers to increase the available phosphorus content in soil,but it can easily lead to soil structure damage,higher costs and increased environmental pollution.Phosphorus solubilizing microorganisms are a class of microorganisms that can convert insoluble phosphate in soil into soluble phosphorus and because of their green and economic,they have been widely used in recent years to increase soluble phosphorus in soil and promote crop growth.Pseudomonas is the most studied genus of phosphorus solubilizing microorganisms due to its strong adaptability.In this study,two strains of Pseudomonas were selected from the laboratory previous screening to investigate their phosphorus solubilization parameters and applied into soil to study their promotion effect on soil phosphorus solubilization and plant growth promotion.Moreover,the potential phosphorus solubilization mechanism of strains were explored through whole-genome sequencing analysis,which provided a foundation for the interpretation and later development and use of phosphorus-soluble microorganisms.The main results of the study are as follows:1.Two phosphorus-soluble strains were selected based on our previous study and they were classified and identified in terms of colony morphology,physiological and biochemical characteristics and 16 S rRNA.The results showed that bacterium 1616X1 was rod-shaped and can grow between 0-5% Na Cl with pH 5-9 range.16 S rRNA sequencing showed99.37% similarity with Pseudomonas putida NBRC 14164;bacterium 1416X3 was long rod-shaped and can grow between 0-3% Na Cl in range of pH 4-8,which 16 S rRNA sequencing showed 99.65% similarity with Pseudomonas fildesensis KG01.The liquid culture test and molybdenum antimony anti-colorimetric method was used to determine the solubilization of insoluble phosphate by both strains,and the results showed that the solubilization of phosphorus by strain 1616X1 was 629.47,1048.99,7.36,25.04 and 52.01mg/L for calcium phosphate,calcium hydrogen phosphate,iron phosphate,aluminum phosphate and calcium phytate,respectively,and the solubilization of phosphorus by strain1416X3 was 690.11,1049.98,8.22,22.57 and 49.33 mg/L for strain 1416X3,respectively.2.The phosphorus solubilization parameters of strain 1616X1 were optimized by single-factor and response surface tests.The results showed that the optimal conditions for strain 1616X1 were glucose 10 g/L,ammonium sulfate 0.17 g/L,initial pH 5.70,inoculum5%,loading volume 75 m L/250 m L,and incubation temperature 23.3°C.Within this conditions,the phosphorus solubilization amount could reach 684.92 mg/L,which was17.26% higher than non-optimization.Moreover,the phosphorus dissolution conditions of strain 1416X3 have been optimized in the laboratory,and the results showed that under the conditions of glucose 15 g/L,ammonium sulfate 0.10 g/L,initial pH 7.50,inoculum 2%,and temperature 28°C,the phosphorus solubilization amount could reach 1005.20 mg/L.3.The results of soil phosphorus solubilization and maize growth promotion showed that the highest soil fast-acting phosphorus content was 11.18 mg/L in the sterilized soil system with strain 1616X1,which increased by 108.19%,and maize plant height,root length,fresh weight,dry weight and total phosphorus increased by 25.89%,77.97%,64.71%,72.23% and70.10%,respectively compared with the uninoculated system;The highest soil fast-acting phosphorus content of 11.38 mg/L was achieved with strain 1416X3 inoculation,which increased by 111.92%,and maize plant height,root length,fresh weight,dry weight and total phosphorus increased by 16.88%,114.04%,80.43%,81.30% and 88.42%,respectively.In the unsterilized soil system,the soil fast-acting phosphorus content was 7.67 mg/L under the action of strain 1616X1 with an increase of 107.21%,and maize plant height,root length,fresh weight,dry weight and total phosphorus increased by 28.22%,14.69%,47.53%,52.20% and 23.09%,respectively,compared with the uninoculation system;while the soil fast-acting phosphorus content under the effect of strain 1416X3 was 8.11 mg/L,an increase of 118.92%,and the maize plant height,root length,fresh weight,dry weight and total phosphorus increased by 33.63%,10.11%,38.42%,46.57% and 28.72%,respectively.The effect of strain inoculation on soil microbial community structure was further analyzed.After8 days inoculation,the inter-rhizosphere soil inoculated with strain 1616X1 was enriched with Pseudomonas and Nocardioides,while Arthrobacter,Sphingomonas and Massilia was decreased;the soil inoculated with strain 1416X3 showed a increasing trend of Arthrobacter,Nocardioides,and Knoellia as well as Bacillus,Gaiella,and Solirubrobacter showed a decreasing trend.4.Whole-genome sequencing analysis of strains showed both strain 1616X1 and 1416X3 contains the genes associated with phospholysis.The genes encoding glucose dehydrogenase(gdh),cofactor pyrroloquinolone quinine(pqqBCDE)and the Pst phosphorus transport system(pstS,pstC,pstA,pstB and phoU),but the genome of strain 1416X3 has more Pst phosphorus transport system genes than strain 1616X1 and contains genes encoding alkaline phosphatase(phoD);In the tricarboxylic acid cycle,pyruvate metabolic pathway,contains a variety of organic acid synthase involved in strain 1616X1 and 1416X3 solubilization of inorganic phosphorus,which encodes organic acid synthase genes are IDH3,aarC,ldhA,maeB,etc.,and the genome of strain 1416X3 encodes more organic acid synthase genes than strain 1616X1;In addition,both strains contain genes encoding In addition,both strains contained genes encoding tryptophan synthase and synthesized indole acetic acid through tryptophan metabolism,but the intermediate products produced and the enzyme classes were different. |
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